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Title: Section-Cutting: A Practical Guide - To the preparation and mounting of sections for the microscope, etc.
Author: Marsh, Sylvester
Language: English
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SECTION-CUTTING:
A PRACTICAL GUIDE
TO THE
PREPARATION AND MOUNTING OF SECTIONS FOR
THE MICROSCOPE,
_SPECIAL PROMINENCE BEING GIVEN TO THE
SUBJECT OF ANIMAL SECTIONS._
BY
DR. SYLVESTER MARSH.
_WITH ILLUSTRATIONS._
REPRINTED FROM THE LONDON EDITION, WITH NOTES AND AN APPENDIX
ON THE SELECTION AND CARE OF SECTION KNIVES.
NEW YORK:
INDUSTRIAL PUBLICATION COMPANY.
1879.
------------------------------------------------------------------------
PREFACE.
If we glance at any of the numerous magazines devoted either wholly or
in part to the subject of microscopy, we shall hardly fail to be struck
with the numerous queries relating to SECTION-CUTTING, which are to be
found in its pages. A simple explanation of this wide-spread want is
afforded by the fact that the use of the microscope has at the present
day extended to (and is still rapidly spreading amongst) vast numbers of
students, who, in many instances, possess neither the leisure nor the
means to refer for information to large and expensive text-books.
Moreover, were they actually to consult such works, they would
practically fail to obtain the information of which they are in need,
for the coveted instruction is to be found in those treatises only in a
scattered and fragmentary form—no work with which we are acquainted
treating of the subject in anything like a detailed manner. To fill this
_vacuum_ in the literature of microscopy the present manualette has been
prepared. Little claim is made to originality, yet the book is by no
means a mere compilation, but the outcome of long and extensive personal
experience in the cutting and mounting of microscopical sections. Every
process described has been put to the test of actual trial, so that its
worth may confidently be depended upon. Many of the little points
insisted upon in the ensuing pages will doubtless to the practised
microscopist appear superfluous or even puerile; but a vivid
recollection of our own early failures and disappointments assures us
that it is just these very _minutiæ_ of detail which will be found most
serviceable in directing and sustaining the faltering footsteps of the
tyro.
ST. HELENS, _September, 1878_.
------------------------------------------------------------------------
PREFACE TO AMERICAN EDITION.
--------------
Dr. Marsh’s book cannot fail to promote the practical study of histology
wherever the English language is spoken, and under ordinary
circumstances the students of this country might have been left to the
use of the London edition. But a careful reading showed that one or two
points which are perfectly clear to British readers might be a source of
doubt and difficulty to students on this side. At the present day in
this country histology is a favorite study with very many private
students who have no access to teachers or large libraries, and for the
purpose of smoothing their path we have added such information as was
obviously desirable. In the hope that these additions will prove of
value, we submit the book to American readers.
NEW YORK, _January, 1879_.
------------------------------------------------------------------------
CONTENTS.
-------
PART I.
Introduction 11
On Cutting Unprepared Vegetable Tissues 12
On Cutting Unprepared Animal Tissues 13
Preparation of Vegetable Tissues 15
Preparation of Animal Tissues 16
Special Methods of Hardening 19
Cutting Hardened Tissues by Hand 19
Microtome 20
Æther Microtome 24
Section-Knife 24
Imbedding in Paraffine for Microtome 26
Employment of Microtome 29
Staining Agents 31
Carmine Staining 32
Mounting Media 36
Mounting in Glycerine 38
Uses of Freezing Microtome 42
Employment of Freezing Microtome 43
Logwood Staining 47
Absolute Alcohol 50
Clove Oil 51
Canada Balsam 51
Mounting in Balsam 53
Finishing the Slide 55
PART II.
Special Methods 57
Bone 57
Brain 59
Cartilage 60
Coffee Berry 62
Fat 63
Hair 63
Horn, etc. 64
Intestine 64
Liver 65
Lung 65
Muscle 66
Orange-peel 67
Ovary 67
Porcupine Quill 67
Potato 67
Rush 68
Skin 68
Spinal Cord 69
Sponge 71
Stomach 71
Tongue 71
Vegetable Ivory 72
Wood 72
Note A, 75
Note B, 75
Note C, 76
Note D, 76
Note E, 76
Note F, 77
Note G, 77
Note H, 93
Note I, 94
Index
------------------------------------------------------------------------
SECTION-CUTTING.
--------------
PART I.
1. _Introduction._—Many of the most interesting objects with which the
microscopist has to deal, cannot be made to reveal their beauty or
minute structure until they have been cut into slices or _sections_, of
such a degree of thinness as to render them transparent, and thus permit
of their examination by transmitted light, with objectives of varying
power. Unfortunately, however, very few of the objects of this class
are, in their natural condition, in a suitable state to be submitted to
this method of procedure. Some are of such a soft and yielding nature
that any attempt to cut them is an utter failure, for in place of a
perfect section being obtained, nothing remains upon the knife but a
mass of diffluent pulp; others, again, are of such density as to resist
the action of any cutting instrument.
It is evident, therefore, that nothing can be done with such refractory
materials until, by subjection to appropriate methods of preparation,
they shall have been reduced to such a consistence as to render them
suitable for cutting. How this is to be accomplished will depend
entirely upon the physical and chemical nature of the substance to be
operated upon. As the various objects differ so widely from each other
in these respects, so must the methods of preparation suitable to each
also vary. It is clear, therefore, that no general directions for
attaining this end can be given which would be of any practical value.
It is possible, however, and very convenient, to arrange the various
objects into groups or classes, to the treatment of each of which
certain general rules are applicable; but there will still remain a
comparatively numerous series of objects whose individual peculiarities
of structure will demand for them correspondingly special methods of
preparation. When such objects come to be spoken of, the particular
treatment most suitable to each will also be noticed.
2. _On Cutting unprepared Vegetable Tissues._—There are some few
substances, however, which may with more or less success be cut into
sections whilst in their natural condition. Such objects are to be found
in the vegetable world in certain kinds of leaves and allied structures,
whilst in the animal kingdom they are principally represented by the
various internal organs of man and the lower animals. Special directions
are given in text-books for the preparation of sections of leaves and
similar substances. For instance, it is recommended to lay the leaf,
etc., on a piece of fine cork, and with a sharp knife to shave off thin
slices, cutting down upon the cork. Another plan is to place the leaf,
etc., between two thin layers of cork, and cut through the mass. No
method, however, is at once so simple and successful as the process of
imbedding in paraffine. To do this, it is necessary to make a paper
mould by twisting a strip of stout writing paper round a ruler, and
turning-in the paper over the end of the ruler. This mould, the height
of which may vary from an inch to an inch and a half, should now be
about half filled with melted paraffine mixture (§ 11), the leaf or
other object plunged into it, and held in position by small forceps till
the paraffine has become sufficiently solidified to yield it a support.
More of the paraffine mixture is now poured in until the specimen is
thoroughly imbedded; the whole is to be put away in a cold place for an
hour or so, when the mass will be found sufficiently firm to be cut with
ease. Sections may be made with a razor kept constantly wetted with
water, or, if the preservation of colour be no object, methylated spirit
may be employed for the purpose. As the subsequent treatment of such
sections in no wise differs from that required by those cut in the
microtome, we shall defer its consideration until that method of section
has been described (§ 12).
3. _On Cutting unprepared Animal Tissues._—For the cutting of fresh
_animal_ tissues several plans may be followed. Thus, if a section of
only very limited area be required, it may be obtained by snipping a
piece off the tissue with a pair of bent scissors, which, for this
purpose, are so made that the blades are _curved on the flat_
(Carpenter). If this be carefully performed it will be found that a
large portion of the section (particularly at the circumference) so
obtained will be sufficiently thin for examination. If a larger section
be desired, an attempt may be made to cut it with a very sharp scalpel
or razor, the blade of which whilst in use must be kept _flooded_ with
water, or spirit, the latter of which is to be preferred. Recourse may
also be had to _Valentin’s_ knife. This consists of two long, narrow
blades, running parallel to each other, the distance at which the blades
are held apart, and which, of course, determines the thickness of the
section, being regulated by means of a fine screw passing through both
blades. A milled head attached to this screw gives a ready means of
opening or closing the blades, so as to bring them to the desired degree
of approximation. The method of using the knife is very simple. After
having “set” the blades at the desired distance apart by means of the
milled head, the tissue to be cut is held in the left hand immersed in a
basin of water. The knife is now steadily and with a rapid motion
_drawn_ through the tissue, care being taken that the cut is made in
such a manner that the blades move from heel to point. By slightly
separating the blades and gently shaking them in the water, the section
at once becomes disengaged. After use, the blades must be thoroughly
dried, when they may be smeared with some oil which does not readily
oxidize. For this purpose, a very suitable oil is that known as
“Rangoon.”[1] Though it has been deemed advisable briefly to describe
the preceding methods of cutting unhardened tissues, it will be found
that for the purposes of the ordinary microscopical student sections so
obtained are of very little value. They are always of very limited
dimensions, seldom of uniform thickness, and often so extremely friable
as to render it very difficult and frequently impossible to submit them
with safety to such further treatment as is necessary to fit them for
being mounted as permanent objects. This method of section-cutting,
however, is not without its uses, for by its means the medical
practitioner is provided with a simple and ready method of roughly
investigating the structure of morbid tissues, whilst to the general
student it furnishes an easy means of making a cursory examination of
certain substances, in order that he may determine whether it be worth
his while to subject them to some of those various processes of
hardening hereafter to be described.
Footnote 1:
Note A.
4. _Preparation of Vegetable Tissues._—Let us now pass to a brief
consideration of the methods usually adopted for preparing the various
objects for easy section. In the case of _vegetable_ tissues, not only
do we, as a rule, find their texture of too great density to be readily
cut in their natural condition, but they also contain much resinous and
starchy matter, of which it is highly desirable to get rid. In order to
do this we first cut the substance (say a stem or root) into small
pieces, which are to be placed in water for three or four days, by which
time all the soluble gummy matters will have disappeared. The pieces are
now transferred to a wide-necked bottle, containing methylated
spirit,[2] which, in the course of a few days, will dissolve out all the
resin, etc. Many kinds of woody tissue are by these processes reduced to
a fit condition for immediate cutting; others, however, are so hard as
to render it necessary to give them another soaking for some hours in
water, to bring them to a sufficient degree of softness to cut easily.
If the wood (as in some few refractory cases will happen) be still too
hard for section, a short immersion in warm, or if necessary, in boiling
water, will not fail effectually to soften it. The treatment of such
members of the vegetable division as require peculiar methods, will be
found described in future pages.
Footnote 2:
Note B.
5. _Preparation of Animal Tissues._—_Animal_ tissues differ from one
another so greatly, both in consistence and in chemical composition, as
well as in their degree of natural hardness, that no general rules can
be given which would be applicable to the preparation of the whole
class. Such as are of any considerable degree of hardness, as horn and
kindred structures, must be treated much in the same manner as the
denser varieties of wood, viz., by more or less prolonged immersion in
water—cold, hot, or boiling. Those which are of extreme hardness, as
bones and teeth, can be cut only by following certain special methods,
full details of which will be found in the Second Part of this work (§
26). Many, and indeed the vast majority of animal tissues, offer a
direct contrast in point of hardness to those we have just been
considering. All the internal organs of the body are, when freshly
removed, of much too soft a nature to permit, when in their unprepared
condition, of easy or perfect cutting. It is upon bringing them to that
critical degree of hardness, which is often so difficult to attain, that
the chief secret of successful section-cutting depends; for unless the
hardening process has been carried up to, but not beyond, a given point,
which varies with different tissues, the operator, however dexterous,
will fail to obtain satisfactory sections. For, if the hardening has
fallen short of this critical point, he is, to some extent, in the same
position as if he were dealing with unhardened tissues; whilst, if this
point has been exceeded, the tissue will have become so brittle as to
crumble before the knife. For the purpose of hardening animal tissues,
the student has at his command two principal agents, namely, alcohol and
chromic acid, each of which possesses advantages of its own, but the use
of each of which is also attended by its own inconveniences. Thus, by
the use of alcohol, there is very much less risk of overhardening the
specimen than if chromic acid had been employed. Alcohol, however,
though a capital indurating agent in some instances, does not answer so
well in many others. Chromic acid is, therefore, to be preferred for
general use. It is, however, a very delicate agent to manage, for unless
the greatest care be taken it is exceedingly likely to overharden
tissues submitted to its action, and when this happens the specimen
becomes utterly useless for cutting, as there is no known means of
removing the extreme brittleness which it has acquired. By taking the
precautions now to be given, this overhardening may generally be
avoided. Let us harden a portion of some viscus, say the kidney, for
instance. Suppose we cut from the organ five or six small pieces (from
half to three-quarters of an inch square, _not larger_). These must be
placed in a mixture of equal parts of methylated spirit and water for
three days, at the end of which period they may be transferred to a
solution of chromic acid, made by dissolving twenty grains of the pure
acid in sixteen ounces of distilled water. The solution should be kept
in a wide-necked bottle furnished with a glass stopper. At the
expiration of seven days, pour off the solution and replace it by fresh.
At the end of another week, carefully examine the immersed tissues, and
by means of a sharp razor see if they have acquired the necessary degree
of hardness to allow of a section of _moderate_ thinness being made. If
so, remove the pieces and put them into a stoppered bottle containing
from six to eight ounces of methylated spirit. If, however, the
hardening be found not to be sufficiently advanced, the chromic acid
solution is to be poured off and again replaced by fresh. It will now be
necessary to examine the tissues at intervals of about two days, until
they are found to be sufficiently hard, when they must be transferred to
the spirit. Under no circumstances, however, should they be permitted to
remain in the chromic acid longer than the end of the third week, and
though they should at this time appear not to have undergone sufficient
induration, yet it will be advisable to transfer them to the methylated
spirit, which in a short time will _safely_ complete the process of
hardening, without any risk being run of the tissue becoming ruinously
brittle. It will be noticed, that when the specimens have been
transferred to spirit, the latter will in a day or two become of a deep
yellow color, whilst a thick flocculent deposit falls to the bottom of
the bottle. The tissues should then be removed, the bottle emptied and
well washed, and, being refilled with clean spirit, the preparations are
again to be replaced. This may occasionally be repeated, until the
spirit becomes and remains perfectly bright and clear. The specimens are
then ready for section.
6. _Special Methods of Hardening._—The brain (§ 27), spinal cord (§ 43),
liver (§ 34), and several other organs, etc., require special methods of
hardening, details of which will be found in the paragraph devoted to
each. In the case of _injected_ preparations, the best plan is to harden
them in alcohol from the outset, beginning with weak spirit, and
gradually increasing the strength as the hardening proceeds. When the
object has been injected with Prussian blue, a few drops of hydrochloric
acid should be added to the alcohol to fix the color.
It may here be observed, that specimens of _morbid tissues_ require, as
a general rule, a shorter immersion in chromic acid solution than
healthy tissues do. A very small degree of overhardening speedily
renders them brittle and useless. They should, therefore, be removed
from the acid medium at the end of ten days or a fortnight, and their
further hardening carried on by means of alcohol.
7. _Cutting Hardened Tissues by Hand._—Our material being now reduced to
a fit condition for cutting, let us proceed to consider the several
methods by which this may be effected. The readiest and most simple
plan, if the piece be large enough, is to hold it in the left hand, and,
having brought the surface to a perfect level by cutting off several
rather thick slices, endeavor to cut a thin section by the aid of a very
sharp razor, the blade of which must be kept well _flooded_ with spirit.
As in the use of _Valentin’s_ knife, so here, great care must be taken
steadily to _draw_ the blade across the tissue, every effort being made
to avoid _pushing_ the knife, else the section will be _torn off_,
instead of being _cut_. Though this method[3] is of very great
importance for many purposes, yet a considerable degree of manipulative
skill is required to enable the operator to obtain anything like perfect
sections by its means, and, unfortunately, this skill is acquired by
very few persons indeed, even after much practice. If the piece which it
is desired to cut be too small to be conveniently held in the hand, it
may be imbedded in paraffine in the manner already described (§ 3). A
very simple imbedding agent, and one of the greatest practical value, is
a strong solution of gum arabic, which, upon being dehydrated either by
ordinary drying or the action of alcohol, soon acquires such a degree of
hardness as to permit it (with the imbedded tissue) to be cut with ease.
As this method of imbedding, however, is most frequently resorted to
where, by its means, special difficulties have to be overcome, a full
description of the process (§ 35) will be deferred until such special
cases come to be spoken of.
Footnote 3:
Note C.
8. _Microtome._—Although the preceding plans may be sufficient to answer
all his requirements, if the student wishes to obtain only one or two
sections of small dimensions, of a given object, if he requires a number
of such sections he will find these methods fail him, for even though by
practice he may have attained to considerable aptitude in the use of the
knife, it will still unquestionably happen that the vast majority of his
sections will be more or less imperfect. If, therefore, it be desired to
procure a number of perfect sections, of equable thickness and large
area, it is absolutely necessary to resort to the use of some form or
other of microtome, or section-cutter. This instrument, in its simplest
form, merely consists of a stout brass tube closed at one end, and being
by the other fixed at right angles into a smooth plate of metal. A plug
or disk of brass, accurately fitting the interior of the tube, is acted
upon by a fine threaded screw piercing the base of the tube, and by
means of which the plug, and any object it may support, can be elevated
at pleasure. The object by this means being made gradually to rise out
of the tube, sections are cut from it by simply gliding a sharp knife
along the smooth cutting-plate, and hence across the specimen. Any
intelligent worker in brass would make an instrument of this kind at a
very small cost, and although perhaps it might lack the finish of an
instrument bought at the optician’s, it would, if accurately made, do
its work as well as the most complicated and expensive. If, however, the
student resolves to purchase a microtome, there are a variety of forms
in the market from which he may choose. A few hints may perhaps be of
service in enabling him to make a judicious selection. At the outset we
may say that unless the student intends to devote himself solely to the
production of sections of wood, etc., he ought not to procure one of
those forms of microtome known as wood section-cutters, in which the
object to be cut is held in position in the tube by means of a binding
screw which pierces its side.[4] Although these machines are all that
can be desired for cutting hard bodies, they are not so suitable for
soft ones. The chief points to be attended to in selecting a microtome
are, (1) that the cutting-plate of the instrument be made of glass, or
in default of this, of very hard metal of the most perfect
smoothness;[5] (2) that the diameter of the tube be neither too large
nor too small—it ought not to be less than 5/8-in., or greater than 1
inch;[6] (3) that the screw, which should be _fine_ and well cut, be
provided with a graduated head; (4) that there be some kind of index by
which fractional portions of a revolution of the screw may be measured;
and (5) that the plug fit the tube of the microtome so accurately that
when melted paraffine, gum, or other imbedding agent be poured into it,
it may not find its way between the plug and side of the tube (§ 18). It
often happens in cutting tissues imbedded in paraffine, that the
pressure of the knife causes the cylinder of the imbedding agent to
twist round in the tube of the machine, and so cause considerable
difficulty and annoyance. This evil is usually met by running a deep
groove across the upper surface of the plug, and into this the paraffine
sinks, and so is prevented from rotating. It will be found, moreover,
that another difficulty of a kindred, though much more serious
character, will frequently be encountered. During section the paraffine
has a tendency not only to rotate, but also to become loosened from the
subjacent plug, and to _rise_ in the tube of the microtome. When this
happens the power to cut sections of uniform thickness has completely
gone, for some will now be found to be many times thicker than others;
in fact, the irregularity in this respect soon becomes so monstrous as
to render it useless to prolong the sitting. In the ordinary run of
microtomes no provision seems to have been made to meet this difficulty,
and for this reason many instruments, of otherwise great merit, have
their efficiency seriously impaired. Fortunately, this imperfection is
easily remedied, all that is required being that the upper surface of
the plug should be furnished with some kind of projection, having at its
summit a table-like expansion, as shown at A in the figure. The
imbedding paraffine, by penetrating beneath and around this, becomes
firmly attached to the plug, and thus all risk of its rising is
effectually avoided. If the student wishes to secure a really
first-class instrument, none can be so confidently recommended as the
freezing microtome of Professor Rutherford. In addition to its being the
best instrument for carrying out the freezing method (§ 18), this
machine is equally effective for cutting tissues imbedded in paraffine,
or any of the other agents used for that purpose; indeed, whatever work
a microtome _can_ do, _this one_ will perform.
Footnote 4:
Note D.
Footnote 5:
Note E.
Footnote 6:
Note F.
[Illustration: SECTION OF MICROTOME-TUBE SHOWING ARRANGEMENT (A) TO
PREVENT “RISE” OF PARAFFINE.]
9. _Æther Microtome._—A word here as to freezing microtomes, where the
agent used is æther. Such as have fallen under our notice have not
answered the expectations we were justly entitled to form of them. That
it is possible to freeze a piece of tissue by their use is undeniable,
but it is, as a rule, at an expenditure of such a quantity of æther
(only the very best of which must be used) as to constitute it a very
expensive proceeding. Another serious disadvantage they possess is, that
if the supply of æther be intermitted for only a very short time, the
already frozen tissue thaws with great and most inconvenient rapidity.
10. _Section-Knife._—Of not less importance than the microtome is the
section-knife, to be used in conjunction with it. How perfect soever the
former, and whatever the dexterity of the operator, unless he be
provided with a suitable and well-made knife, he will never succeed in
obtaining satisfactory results. As to the most desirable _size_ of the
knife, much difference of opinion seems to exist, section knives varying
in this respect from a blade of extreme shortness to one which fell
under our observation, in which the portentous length of _thirteen_
inches was attained. What advantages were to be expected by prolonging
the blade to this extravagant length, must remain an inscrutable mystery
to all save its designer. Concerning the _shape_ of the knife, it is
frequently advised that the surface which has to glide along the
cutting-plate of the microtome should be ground _flat_. A most
unsuitable arrangement, as a very little actual experience of
section-cutting will speedily demonstrate. After many unsuccessful
attempts to obtain a really good and reliable section-knife, we
determined to have one specially made, which, as it has proved
everything that could be desired, merits a brief description. It is of
the utmost importance that the blade be made of good and well-tempered
steel, not only that it may be capable of receiving an edge of the most
exquisite keenness, but also that it may _retain it_. The knife of which
we speak (and which was made by Mr. Gardner, of South Bridge, Edinburgh)
is furnished with a blade _four inches_ long, and 7/8-inch broad, set
into a square handle of boxwood, also four inches in length. The
thickness of the blade at the back is not quite 1/4-inch, while _both_
of its surfaces are slightly hollow ground. It is essentially necessary
that the back and edge of the blade be strictly parallel to each other,
otherwise the knife, when in use, will have such a tendency to tilt over
as to render its management extremely difficult. It is very easy to
discover if this condition be fulfilled, for if on carefully laying the
flat of the blade upon a piece of level glass, every portion of both
back and edge are found to be in close contact with it, the knife may in
this respect be considered perfect. Every student who aspires to be a
successful section-cutter should provide himself with a good Turkey
oilstone, _and learn to use it_. He should also possess a razor strop,
as it will be in constant requisition.[7] It may here be remarked that
though _razors_, as a rule, are unsuitable for use with the microtome
from want of uniformity in the thickness of their blades, yet, if only a
small object is to be cut—for instance, a thin root or stem—very good
results may be obtained from their use, especially if one of the
old-fashioned make, having a thick back and slightly _concave_ surfaces,
be employed.
11. _Imbedding in Paraffine for Microtome._—Having described at some
length the various instruments necessary for section-cutting, we will
now consider how they are to be used. Let us endeavor to cut some
sections—say of a piece of kidney—and in so doing we will adopt the
“paraffine” method of imbedding. Ordinary paraffine, however, when used
alone, is rather too hard for our purpose. In order, therefore, to bring
it to a suitable consistence, it must be mixed with one-fifth its weight
of common unsalted lard, a gentle heat applied, and the two thoroughly
stirred together. A quantity of this should be prepared, so that it may
always be ready when wanted—it is very conveniently kept in an ointment
pot or preserve jar, the top of the latter being well covered, to keep
out the dust. When it is intended to use this mixture for the purpose of
imbedding, only just about the quantity required should be melted; for
in doing this it is advisable to use as low a degree of heat as
possible, not only to prevent injury to the tissue to be imbedded, but
also that the paraffine when cooling may not undergo such an amount of
contraction as to cause it to shrink from the sides of the
microtome-tube. It is therefore a good plan to effect the melting in a
water-bath, a simple kind of which, something after the fashion of a
glue-pot, would be made for a few pence by any tinman.
Footnote 7:
See Note G.
The kidney which we are about to cut has, of course, gone through the
process of hardening already described (§ 5), and is now preserved in
spirit. A small piece, say half an inch square, is selected, removed
with forceps, and placed on a bit of blotting paper, when the surface of
the tissue will rapidly become dry (_only the surface_ must be allowed
to dry). It is the usual plan now to proceed at once to imbed it in the
melted paraffine. This is a most undesirable step, and gives rise at a
later stage of our proceedings to a great amount of trouble and
annoyance, for after sections have been cut from a tissue so imbedded it
will be found that portions of paraffine adhere to their edges with such
tenacity that in the case of many of them there is no effectual method
of removing the paraffine, short of soaking the sections in warm æther;
a very objectionable proceeding, for though the æther will undoubtedly
remove the paraffine, it will also dissolve out any fatty matters which
the section itself may naturally contain. All this annoyance may be
prevented by subjecting the tissue to a simple preparatory treatment
before it is imbedded in the paraffine. For this purpose prepare a very
_weak_ solution of gum arabic in water—twenty grains to the ounce. Into
this, by means of the forceps, dip for a few moments the already
_surface-dried_ tissue, taking special care not to squeeze it, or the
pressure will cause the spirit from its interior to remoisten the
surface, which would prevent the gum from adhering. We shall see the
value of this a little later on. Remove the tissue from the solution on
to blotting paper, when the superfluous gum will speedily drain off, and
in two or three minutes the _surface_ will have become quite glazy and
dry. Having melted some paraffine mixture in the water-bath, the tissue
held in the forceps must be plunged for an instant into the heated
liquid and immediately withdrawn, when the crust of paraffine with which
it is enveloped will promptly harden. Whilst this is taking place we may
make ready the microtome. Having by means of the milled head or handle
depressed the plug in the tube so as to leave a free opening about an
inch deep at its upper end, we must pour in the melted paraffine, which
by this time will have become a little cooler, until the cavity be about
half filled. The prepared tissue must now be introduced, care being
taken to place it in such a position that the sections may be cut in the
desired direction. The tissue must, if necessary, be held in position
with forceps or a needle point, till the imbedding material becomes hard
enough to give it due support. It is here to be remembered that it will
not be advisable to place the tissue in the centre of the tube—it will
be much more easily cut if placed rather nearer to that edge of the tube
which is situated next the operator in the act of cutting. More
paraffine is to be slowly added, until the tissue is completely covered;
even after this still more should be added, for it will be found that in
cooling the paraffine shrinks so as to leave a cup-shaped depression in
its centre, whereby portions of the tissue which were previously covered
are again laid bare. The best method of preventing this is to use the
paraffine at as low a temperature as possible, and to use plenty of it.
The microtome, with its contents, must now be removed to a cool place,
when the paraffine will soon become solidified. Whilst this is being
accomplished we may make our further preparations. The first thing we
require will be a large basin, full of freshly-filtered water, and
provided with a cover. A small beaker of methylated spirit, with a
dipping rod or pipette, will also be necessary. We must now see that the
section-knife is in thorough order, to ensure which it will be advisable
to give it a few turns on the strop. An ordinary razor will also be of
service.
12. _Employment of Microtome._—The paraffine being sufficiently hard, we
will clamp the microtome on to the table, and seat ourselves on a chair
of convenient height before it. To our right stand the basin of water,
razor, and section-knife; the beaker of spirit to the left, and a cloth
on our knee. A few turns of the microtome screw having brought the
paraffine to the surface, a thick slice is to be cut off, and this
repeated until the imbedded tissue comes into view. This preliminary
work had best be done with the razor, as it is needless to subject our
section-knife to unnecessary wear and tear. By a fractional revolution
of the screw the tissue is now slightly elevated, and with the pipette
held in the left hand, a large drop of spirit is to be let fall upon its
surface. The section-knife, grasped firmly but lightly in the right
hand, is to be laid flat upon the cutting-plate of the machine, so as to
occupy the diagonal position shown in the figure. Two fingers of the
left hand are now laid gently upon the back of the blade, so as to give
it an equable support, whilst the knife with a rapid motion is pushed in
the combined direction of _forwards_ and to the _left_, so that the
blade in cutting the tissue will pass through it from point to heel.
Thus it will be observed that the stroke of the knife is _from_ the
operator—a far easier and more effective mode of cutting than the
reverse plan. The blade of the knife, having the section just cut,
either floating in a small pool of spirit on its surface or adhering
thereto, must now be immersed in the basin of water, when by a little
very gentle agitation of the knife the section will be floated off. And
now we shall find the great practical value of immersing the tissue in
gum before imbedding, for no sooner is the section disengaged from the
knife than the thin film of gum which separates the paraffine from it
becomes dissolved, and the section will be observed gradually to subside
to the bottom, leaving the paraffine floating upon the surface. After
carefully wiping the knife from all shreds of paraffine, the microtome
screw must again be partially revolved, more spirit applied to the
tissue, and another section being cut, it must be transferred to the
water as before, and so on, until a sufficient number of sections have
been obtained. As to how thin the sections should be cut, no general
directions can be given; each case must be regulated by its own
conditions. The denser the tissue, the thinner should the section be;
whilst certain substances of loose and spongy texture do not require the
sections to be particularly thin—it may be said, however, in a general
way that sections, and especially animal ones, _cannot be cut too thin_
so long as they remain perfect and entire. If Professor Rutherford’s
microtome (as made by Gardner) be employed, the head of the screw will
be found to be graduated into divisions of slightly unequal value; the
sections will therefore be marked by corresponding variations of
thickness, so that amongst a number cut, there must be many of the exact
thickness to meet the requirements of any individual case.
[Illustration: DIAGRAM SHOWING DIAGONAL POSITION OF KNIFE IN
COMMENCING TO MAKE A SECTION.]
13. _Staining Agents._—Before proceeding to mount the sections which
have just been cut, it will be very advisable that they should be
submitted to the action of some staining fluid, in order to render more
clear and distinct their minute structure. Organic substances possess
the property of being able to absorb various colouring matters from
their solution, and to incorporate such colour into their own texture.
This power of attraction is not, however, possessed by all substances
indiscriminately, or to an equal extent. Some possess it in a high
degree, while others appear to be nearly, if not entirely, devoid of
such power. Hence it follows, that if we immerse an organic tissue (one
of our sections, for instance) of complex structure, in a suitable
staining fluid, the tissue will not become stained in an even and
uniform manner throughout, but the several portions of it will receive
varying depths of colour in accordance with the varying attractive power
of its several constituents. By this means we are enabled in stained
sections to discriminate by their difference of shade, minute and
delicate structures, which in the unstained condition it would be
difficult and often impossible to differentiate. For the purpose of
section-staining there are many agents in use, the most generally
suitable being carmine, logwood (§ 19), and aniline blue (§ 27); whilst
for special purposes chloride of gold (§ 28), pyrogallate of iron (§
28), and several others are all of much value.
14. _Carmine Staining._—In the case of animal sections, carmine is, as a
rule, to be selected, giving as it does most satisfactory and beautiful
results. Tissues may be stained with carmine by two different plans: in
the first, a strong solution is used, and the tissue subjected to its
action for a very short period only, whilst in the latter only very weak
solutions are employed, the time of immersion being considerably
prolonged. The rapid method, however, is not to be recommended, for the
strong carmine acts so powerfully upon the tissue as to give the various
elements comprising it no time, as it were, to exercise their power of
quantitive selection, but involves the whole in one uniform degree of
shadeless colour. By adopting the gradual method much better results are
obtained, each portion of the tissue being now at liberty to acquire its
own particular shade. Amongst the various formulæ for the preparation of
carmine fluid, none can be so safely followed as that devised by Dr.
Lionel Beale. It runs thus:—Place ten grains of the finest carmine in a
test tube, add thirty minims of strong liquor ammonia, boil, add two
ounces of distilled water, and filter; then add two ounces of glycerine,
and half an ounce of rectified spirit—this solution ought to be kept in
a well stoppered bottle. The best vessels in which to stain sections are
small jars of white porcelain, capable of holding about two fluid
ounces, and furnished with lids—they are much preferable to beakers or
watch glasses, for owing to the white background which they afford it is
very easy to watch how the staining is proceeding. The carmine solution
which we have just described is both too strong and of too great density
to be used in its pure state. It will, therefore, require to be diluted
with distilled water before use—the most useful degree of dilution being
attained by adding one part of stain to seven of water. Sections may be
placed in this solution for twenty-four hours, in which time they will
usually be found to have acquired a sufficient depth of colour. If,
however, the tissue be unusually difficult to stain, the time of
immersion may be doubled, or still further prolonged, without detriment
to the section.
[Illustration: SECTION SPOON.]
Having prepared and filtered some of this dilute solution, say an ounce,
let us proceed to stain with it those sections which we left in the
basin of water (§ 12). Here we are at once met by a practical
difficulty. How are the sections to be transferred from one vessel to
the other? This is ordinarily effected by means of a soft camel’s-hair
pencil. It is a method, however, open to grave objections, for the
sections so curl around the brush, and get entangled amidst its hairs,
that, notwithstanding every care, valuable sections not unfrequently
become torn during transit. Every difficulty at once vanishes if we
substitute for the brush a small implement, which any one can readily
make for himself. All that is necessary is to take a strip of
German-silver, or copper, of the thickness of stout cardboard, and about
seven inches in length by five-eighths of an inch in breadth. The sharp
angles are to be filed off and the edges carefully smoothed, whilst at a
distance of five-eighths of an inch from each extremity the end must be
turned up so as to form an angle of about 35°. One end must be left
plain, whilst the other, with the aid of a punch or drill, is to be
pierced with five holes about the thickness of a stocking needle[8] (see
Figure). If we now dip the perforated end of this spoon into the water
containing the sections, and gently agitate it, the sections will rise
from the bottom and float about. The spoon is now brought under one of
them, and being steadily lifted up the water flows downwards through its
apertures, and the section smoothly spreading itself out upon the spoon,
may be gently lifted out of the water, and on the spoon being dipped
into the staining fluid the section at once floats off. By this simple
means sections, however large, thin or delicate, may with ease be
conveyed from one fluid to another, with the utmost certainty of their
not being injured during the process. The sections having been in the
carmine fluid for about twenty-four hours, as much of the liquor as is
possible must be gently poured off, and its place supplied by a
freshly-filtered mixture of five drops of glacial-acetic acid to one
ounce of water, when in a few moments the carmine will become
permanently _fixed_ in the tissue, and the process of staining be
complete.
Footnote 8:
Dr. Klein describes a kind of “lifter,” made by bending some
German-silver wire, but as no drawing accompanies his description, it
is not easy to form a clear idea as to the form of this instrument. In
the recent and philosophical work of Schäfer, a lifter is figured,
which consists of a wire stem, having attached to its end a spade-like
blade. It will be observed that the spoon described in the text
differs from this lifter in having one end perforated, and in this
consists the real value of the implement.
15. _Mounting Media._—The further treatment of the stained sections will
entirely depend upon the nature of the medium in which it is intended to
mount them. There are a variety of fluids in use for this purpose, the
principal being dilute alcohol (§ 26), dammar, or Canada balsam (§
22-23), and glycerine. These, however, cannot be used indiscriminately,
each possessing certain special properties which render it suitable for
use with particular classes of objects only. Thus, weak spirit, having
no tendency to increase the transparency of objects, can advantageously
be used with such only as are already perfectly transparent. It is also
more suitable for the preservation of vegetable tissues (when the
retention of colour is no object) than animal, since with the latter it
has a tendency after a while to cause a kind of granular disintegration,
which ultimately destroys much of the usefulness of the preparation.
Dammar and Canada balsam, on the other hand, possess very great
refractive power, so that they are of great service in mounting objects
which require their transparency to be much increased. For this reason
they are not well adapted to the preservation of very delicate or
transparent tissues (unless previously stained), the minute details of
which become almost entirely obliterated when mounted in them. The chief
advantage possessed by these resinous media is, that tissues mounted in
them undergo no alteration, even after the lapse of many years.
Glycerine, in respect of its clarifying powers, occupies an intermediate
position between spirit and balsam, being much more refractive than the
former, infinitely less so than the latter. It is, therefore, of very
great value for the preservation of such tissues as possess a medium
degree of transparency, and which would become obscured if mounted in
spirit, or have their outlines rendered indistinct if preserved in
balsam. It is of the utmost value for mounting unstained anatomical
sections which, when put up in this medium, reveal such minute details
of structure as would readily have escaped observation had any other
agent been employed. It may also be used with stained sections, but in
this case the sections should be of extreme thinness, otherwise the
refractive power of the glycerine will be insufficient to render them
thoroughly transparent. The great drawback to the use of glycerine is
the extreme difficulty experienced in preventing its escape from beneath
the covering glass, for it unfortunately possesses such great
penetrating power that no cement hitherto devised can be thoroughly
depended upon for withstanding its solvent action for any considerable
length of time.[9] Attention to the instructions presently to be given
(§ 16) will, however, reduce this risk of leakage to a minimum. In the
use of glycerine Dr. Carpenter’s caution must ever be borne in mind,
viz., that, as carbonate of lime is in time dissolved by glycerine, this
agent ought never to be employed for the preservation of objects
containing such salt.
Footnote 9:
Note H.
16. _Mounting in Glycerine._—To illustrate the method of using this
medium we will mount our present sections in glycerine. In the first
place we shall require a deep watch-glass, which is to be half filled
with glycerine diluted with an equal amount of distilled water. By means
of the spoon, one or more sections may be transferred into this, either
directly from the acetic acid solution (§ 14), or if, since cutting,
they have been preserved in spirit, they should first undergo a short
immersion in a large vessel full of water. The watch-glass should now be
covered with an inverted wine-glass, and put away for some hours, in
order that the sections may become thoroughly saturated with the dilute
glycerine. When this has been accomplished, a slide must be cleaned, and
one of the sections, with the aid of the _unpierced_ end of the spoon,
be transferred to its centre.[10] As the kind of section with which we
are now dealing is, or ought to be, of extreme thinness, no cell (§ 26)
is necessary. After tilting up one end of the slide, so as to drain off
as much of the weak glycerine as possible, a drop of Price’s best
glycerine must, with a glass rod or pipette, be allowed to fall gently
upon the section, so as to avoid the formation of air-bubbles. If any of
these, however, should be produced, they must be removed with the point
of a needle set in a wooden handle,[11] and the slide then covered with
a small bell-glass (or wine-glass). A circular cover is now to be
cleaned with a soft handkerchief, and after gently blowing from it any
adhering fibres of lint, etc., it will be advisable to hold the side of
the glass which is to come into contact with the preparation close to
the mouth, and breathe upon it, so as to cover it with moisture. The
cover held between the thumb and forefinger of the left hand must now be
applied by its edge near to the margin of the preparation, and the
surface of the cover directed in an inclined manner over it. Beneath the
overhanging edge of the cover the point of the needle, held in the right
hand, is now to be inserted (see Figure). By gently lowering the needle,
the cover will come into gradual contact with the slide, driving before
it a minute wave of glycerine, in which any air-bubbles that may have
become developed are usually carried off. A very considerable degree of
tact, however, is required to perform this little operation, simple as
it may appear, for the retreating wave of glycerine not unfrequently
floats out the section, either wholly or partially, from beneath the
cover. Air-bubbles, also (the _bêtes noires_ of this process), are
exceedingly likely to arise. When this happens the best plan to adopt
is, by means of the needle point, gently to raise and remove the cover,
apply another drop of glycerine to the section, and cover _with a fresh
piece of thin glass_. It will now be necessary to remove any superfluous
glycerine which may have collected around and near the cover. The great
bulk must be wiped away by means of a camel’s-hair pencil, slightly
wetted between the lips, any remaining stickiness being removed with a
bit of blotting paper which has been slightly damped. With a very small
camel’s-hair pencil, charged with solution of gelatine, a ring must be
made round the margin of the cover, of sufficient breadth to take in a
small tract of both cover and slide. As this cement is perfectly
miscible with glycerine, it readily unites with any of that fluid which
may ooze from beneath the cover, and which, in the case of any of the
ordinary varnishes, would act as a fatal obstacle to perfect adhesion.
To make the cement, take half an ounce of Nelson’s opaque gelatine, put
in a small beaker, add sufficient cold water to cover it, and allow the
mixture to remain until the gelatine has become thoroughly soaked. The
water is now poured off, and heat applied until the gelatine becomes
fluid, when three drops of creosote should be well stirred in, and the
fluid mixture transferred to a small bottle to solidify. Before use,
this compound must be rendered liquid by immersing the bottle containing
it in a cup of warm water. When the ring of gelatine has become quite
set and dry (which will not take long), every trace of glycerine must be
carefully removed from the cover and its neighborhood, by gently
swabbing these parts with a large camel’s-hair pencil dipped in
methylated spirit. After drying the slide a ring of Bell’s microscopical
cement may be applied over the gelatine, and, when this is dry, another
coat is to be laid on. If it be desired to give the slide a neat and
tasteful appearance, it is a very easy matter, by means of the
turn-table, to lay on a final ring of Brunswick black or white zinc
cement (§ 24). Every care has now been taken to render our preparation
permanent; but, to make assurance doubly sure, it will be well to follow
Dr. Carpenter’s advice, and, every year or so, to lay on a thin coating
of good gold-size.[12]
Footnote 10:
The appearance of a slide is vastly improved if the preparation be
placed _exactly_ in its centre. This may readily be done in the
following manner:—Take some very finely-powdered Prussian blue, and
rub it up in a mortar with a little of the weak gum solution (§ 11),
so as to form a thin blue pigment. A quantity of this should be made,
so as always to be at hand. A slide having been cleaned, the _best
surface_ is to be selected, and on the _reverse_ side, by means of the
self-centring turn-table, a small circle is to be drawn with a
camel’s-hair pencil, charged with the pigment. In the centre of this
ring, but on the opposite side of the slide, the section is to be
placed, when it of course will occupy a position exactly central. When
the slide comes to be finished, the blue ring may easily be removed
with a wet rag.
Footnote 11:
A _crochet-needle_ holder made of bone, and which may be bought at
the smallware dealers’ for about sixpence, makes an admirable handle
for microscopical needles. At one extremity there is a small cavity,
closed with a cap, for the storage of reserve needles, whilst the
other end terminates in a metal tip, provided with a crucial slit and
central perforation for the reception of the needle in actual use, and
so arranged that, by means of a small screw-nut, needles of various
sizes may be firmly held in position.
Footnote 12:
If square covers be employed, they may be fixed to the slide by a
simple method much in vogue in Germany. A thin wax taper is to be
lighted, and being partially inverted for a few seconds, the wax
surrounding the wick will become melted. After the slide has been
freed from excess of glycerine, a drop of this heated wax is allowed
to fall upon each corner of the cover, and a line of the melted wax
run along the margins of the cover between these points, so as
perfectly to surround it. If a good coat of white zinc cement be
subsequently laid over the wax a very durable, and not unornamental,
line of union will have been formed.
[Illustration: METHOD OF APPLYING COVER.]
17. _Use of Freezing Microtome._—Our preceding consideration of the
method of employing the microtome in conjunction with paraffine as an
imbedding agent (§ 11), will have formed a very suitable introduction to
the study of the somewhat more complicated process of imbedding the
tissue in gum, for section in the freezing microtome. This method is of
the utmost value to the practical histologist, for by its means he is
enabled with ease to possess himself of perfect sections of several
structures, the cutting of which, before the introduction of this
process, was always a matter of difficulty and anxiety. The freezing
microtome is especially valuable for the section of such substances as
from their extreme delicacy are liable to be injured by being imbedded
in paraffine—for instance, the delicate villi of the intestines becomes
very frequently, by the use of paraffine, denuded of their epithelium,
and the villi themselves not seldom become torn off or otherwise
damaged. The great value of the method is also very well seen in the
treatment of those tissues which, like the lung, are of such loose and
spongy texture as to offer insufficient resistance to the knife unless
their interstices have previously been filled up with some solid yet
easily cut material. As the space at our command is strictly limited, we
are precluded from entering as fully into this branch of section-cutting
as the importance of the subject demands and our own inclination would
lead us. To those who wish to become thoroughly conversant with the full
value of this method we cannot do better than recommend the perusal of
Professor Rutherford’s _Practical Histology_, 2d edition, than which, on
the whole subject of physiological microscopy, no treatise with which we
are acquainted is at once so plain, practical, and profound.
18. _Employment of Freezing Microtome._—A very suitable object with
which to demonstrate the method of using this form of microtome will be
afforded us by a portion of intestine, say of the ileum of a cat or dog.
Suppose we have some of this in methylated spirit—let us select a piece
of about half an inch in length. Our first care will be to deprive this
of its spirit; for so long as the tissue remains impregnated with
alcohol it would, of course, be impossible to freeze it. We will,
therefore, throw it into a large basinful of water, and leave it there
for twenty-four hours, during which time it would be as well to change
the water once or twice. We shall now require a strong solution of gum.
This, which should have been made some time previously, may be prepared
by placing a quantity, say three or four ounces, of ordinary gum arabic
in a glass beaker, and adding sufficient water to cover it—the mixture
must be stirred occasionally with a glass rod until solution has taken
place, which will be in a few days. If _necessary_ a little more water
may be added, but so long as the gum will pour from vessel to vessel, it
cannot well be made too strong. Mucilage, by keeping, is very apt to
become sour and mouldy—this may be prevented by adding to each ounce of
the water with which it is prepared about half a grain of salicylic
acid. We now pour some of this mucilage into a small vessel—an egg cup
will answer very well—and into it transfer the piece of ileum from the
water. Here we must allow it to remain for a time sufficient to permit
of its becoming thoroughly saturated with the gum, for which purpose
some hours will be necessary. When this soaking has been accomplished we
will prepare the microtome, which we will assume to be Rutherford’s. In
the first place it will be necessary to remove the plug—which is to be
done by turning the handle connected with the screw until the plug rises
so high in its tube that it may be grasped with the fingers and removed,
when it is to be well smeared all over with sperm oil and replaced. This
is done to prevent any unpleasant adhesions taking place whilst the
freezing is going on. We must next depress the plug, so as to convert
the upper part of the tube into a kind of “well” of sufficient depth to
hold our specimen. It will now be very advisable to look carefully into
this _well_ and observe whether the plug fits accurately into the tube
(§ 8) for if there be any interval between the two it will give rise to
much subsequent annoyance, as the gum penetrating this interstice will
there become firmly frozen into irregular patches, which will so
interfere with the even gliding of the plug within its tube as to cause
the former to ascend in such an irregular and jerky manner as to be
utterly destructive of all accuracy in the cutting. If this defect be
observed, it may be at once remedied by dropping a small quantity of
gently heated paraffine into the _well_, which will effectually close up
any fissures. The microtome, by means of its clamping arrangement, must
now be firmly attached to the table, and a suitable vessel be placed on
the floor beneath it, so that it may catch the water which will issue
from the waste-pipe of the apparatus. The next requirement is a supply
of block ice and finely-powdered salt. A lump of the ice must be wrapped
in a towel, and crushed into small pieces; these, by means of a large
mortar, are to be further reduced to a very _fine powder_. Any attempt
to hurry over this troublesome part of the operation will lead to future
disappointment, for unless the ice be used in a very fine powder great
delay (at least) in the freezing will be the result. With the aid of a
small spoon the ice and salt are in alternate spoonsful to be conveyed
into the freezing-box of the machine, great care being taken that the
cavity under the cutting-plate and around the tube be thoroughly packed,
after which the uncovered portion of the box should also be well filled.
The _well_ is now to be filled with the strong gum to within a little
distance of its top, and a piece of sheet gutta-percha (such as shoe
soles are made of) being applied over the well, and kept in position by
a weight, we must wait until the freezing commences. In a short time we
shall notice that the gum has acquired a thick muddy appearance. The
tissue must now, by means of the forceps, be transferred to the well,
and there placed in such a position that the sections, when cut, shall
run in the desired direction. After more gum has, if necessary, been
added, so as completely to cover the tissue, the well is again to be
covered, and attention given to the freezing-box. As the mixture which
this contains becomes melted, it must constantly be renewed, care being
at the same time taken that the mouth of the discharge-pipe be kept
quite free, otherwise water accumulating in the box, the freezing
mixture will degenerate into a useless puddle. When the gum becomes
sufficiently hard to cut, this must be done much in the same manner as
if paraffine had been used (§ 12). In this case, however, no fluid will
be required, or must be used, to wet the knife with, and especial care
must be taken that in disengaging the sections from the knife into the
water they be not torn. These sections often adhere very tenaciously to
the blade, but if a little patience be exercised the water will soon
float them off in safety—much more safely than if any attempt be made to
liberate them prematurely. There is one circumstance connected with the
use of the freezing microtome which is rather annoying. The moisture of
the breath and atmosphere is apt to become condensed on the
cutting-plate, and here, mixed with accidental smears of gum, it becomes
frozen into a jagged and irregular sheet of ice, which not only
seriously interferes with the smooth play of the knife, but also
constitutes a real peril to its edge. As this evil cannot be avoided,
all we can do is, by constant wiping, to keep the cutting-plate clean
and free from this accumulation. This is best done with a bit of soft
rag _just moistened_ with spirit, but this must not come into contact
with any portion of the cylinder of frozen gum, else it will instantly
thaw it. When using the freezing microtome it is always advisable to
wear an apron, otherwise our clothes may receive considerable damage
from the constant splashing of the salt water, as it falls from the
waste-pipe into the vessel beneath it. After use, the microtome must be
well washed in plenty of cold water till every trace of salt be removed,
for if any of this remain it will quickly corrode the brass-work of the
instrument. The plug and screw, as also the section-knife, should be
well smeared with Rangoon oil before the machine is put away.
19. _Logwood Staining._—The employment of logwood as a staining agent is
now becoming very general. It acts much in the same manner as carmine,
but the violet color which it produces is by many thought to be of a
more soft and agreeable character than that due to the action of
carmine. A valuable and very convenient property also which it possesses
is that it stains tissues very rapidly, and this without interfering
with that differential kind of coloration (§ 14) upon which the chief
value of all staining processes depends. A simple method of preparing
the logwood fluid is to mix an aqueous solution of extract of logwood
with a solution of alum (1 to 8) till the deep impure red colour has
become violet, and then to filter the mixture (Frey). This will stain
sections in about half an hour. This stain, though here mentioned for
the ease with which it may be made is, as a rule, very inferior to a
fluid prepared directly from hæmatoxylon, the alkaloid or active
principle of logwood. As, however, it is difficult and troublesome to
make the solution in this manner,[13] it will be advisable for the
student to purchase, ready prepared, such small quantity of the dye as
he may require. Small bottles may be obtained for a few pence of Mr.
Martindale, 10, New Cavendish Street, London, and from repeated trials
of this solution we can recommend it as producing excellent results. It
is a very strong fluid, and requires to be diluted before use. The
degree to which the dilution must be carried cannot, however, be very
accurately indicated, for all staining fluids of this nature possess the
very undesirable property of becoming decomposed by age. After the fluid
has been kept for some time, a portion of the colouring matter is thrown
out of solution, and becomes deposited upon the sides and bottom of the
vessel in which it is contained, hence the older the preparation, the
weaker it will have become. As the time required for staining with
logwood is but short, it is desirable that all the sections should begin
to be submitted to its action at the same time, otherwise some will
become more deeply stained than others. A good plan is to fill a small
porcelain jar (§ 14) with filtered water, and into this transfer the
sections. Whilst they are settling well down to the bottom, a mixture
must be prepared of half a drachm of Martindale’s solution (fresh) to
one ounce of distilled water, and everything got in readiness for its
immediate filtration. The water is now very gently to be poured off the
sections, and if care be exercised this may be done in such a manner as
to leave them undisturbed at the bottom, after removing almost every
drop of water. The diluted logwood fluid must now be _immediately_
filtered upon the sections, so that they may run no risk of becoming
dry. In the present instance the staining may be allowed to proceed for
about thirty minutes, and this will be found a convenient time for the
immersion of the general run of animal sections. If the logwood fluid be
not quite fresh, either a little more of it will have to be added to the
water, or the time of immersion must be prolonged until the desired
depth of color has been produced. It is well whilst the staining is
going on gently to shake the vessel occasionally, so that the sections
may not remain in a heap at the bottom, but all be as fully as possible
exposed to the action of the dye. When the staining is judged to be
complete, the logwood solution must be gently poured off, leaving the
stained sections at the bottom of the jar, when they should be quickly
covered with methylated spirit, which will _fix_ the colour. We shall
now be able to see if the coloration obtained be perfectly satisfactory.
If not deep enough, it is very easy again to submit them to the action
of the dye for a few minutes longer. If on the other hand, and as more
frequently happens, the coloration should be too deep, the excess of
colour may readily be removed by transferring the sections for a short
time into some diluted acetic acid prepared by adding five drops of the
glacial acid to an ounce of water. The action of this should be
carefully watched, and when the colour has been reduced to the desired
tint the sections may be retransferred to the methylated spirit.
Footnote 13:
Should the student, however, determine to prepare this solution for
himself, he will find a good formula for the purpose in Schäfer’s
“Practical Histology,” p 176. Note I.
20. _Absolute Alcohol._—As we purpose mounting the sections which have
just been stained, in Canada balsam, we will briefly consider the
preliminary treatment to which they must be submitted before this can be
effected. The object of this is to abstract from the tissue all its
water, for if any moisture be permitted to remain in the section it
will, when mounted in balsam, become obscured and surrounded by a kind
of opalescent halo, due to the imperfect penetration of the balsam into
the only partially dehydrated tissue. The old-fashioned plan of
dehydration was simple exposure to the air. The method now generally
adopted is to bring about the same result by means of absolute alcohol.
This fluid has such a strong affinity for water that tissues submitted
to its influence are rapidly and effectually deprived of any water they
may contain. Absolute alcohol in small quantity may be obtained from the
druggist at about sixpence per ounce. It will be necessary for the
student to provide himself with a little of this agent, say about two
ounces, the method of using which will very shortly be explained.
Absolute alcohol must be kept in a bottle with a very accurately-fitting
stopper, in order to prevent its absorbing moisture from the air. For
our purpose such a bottle, having a neck _as wide as possible_, is to be
selected.
21. _Clove Oil._—After being thoroughly dehydrated the sections may, in
special instances (§ 48), be at once mounted in balsam; but, as a
general rule, it will be found necessary (particularly in the case of
animal sections) to treat them with some clarifying agent, in order to
remove the cloudiness and opacity which is (in part) due to their
previous immersion in alcohol. For this purpose turpentine, or any of
the essential oils, may be used: of these, oil of cloves is to be
specially recommended. It is rather expensive, ranging from sixpence to
one shilling per ounce: but, as a drop or two will be sufficient for
preparing each slide, only a small quantity—say half an ounce or an
ounce—need be procured. The most convenient vessel in which to keep the
oil is one of the small test bottles used by watchmakers. These bottles
are provided with a glass cap to exclude dust, and the stopper is
prolonged into a glass rod, which dips into the bottle. The use of this
rod and the method of employing the oil will be explained shortly.
22. _Canada Balsam_, as ordinarily met with, is a thick resinous balm of
great viscidity, but readily rendered perfectly fluid by the application
of heat. Formerly, sections were mounted in this medium in its pure
state, but owing to the annoyance which was so constantly being
experienced from the tenacity with which intruding air-bubbles were held
by the viscous medium, this plan of mounting is rapidly falling out of
use.[14] It is now usual to employ the balsam in a diluted condition,
the two chief diluents being chloroform and benzole. As balsam, however,
often contains more or less moisture, it is desirable to drive this off
before adding the diluent. A very convenient way of doing so is to
expose some pure balsam to the heat of a cool oven for several hours,
when the balsam will be found to have assumed a hard, vitreous
character. It should now be broken into small pieces, these put into a
bottle, and some methylated chloroform added, which in a little while
will completely dissolve the hardened balsam. More chloroform is then to
be added, until a solution is obtained sufficiently thin to run through
filtering-paper. A glass spirit lamp must now be procured, having a
capacity of about two ounces, and provided with a cap. Into the
wick-holder of this (which must be made of porcelain) a hollow glass
tube is to be so fitted that its end dips into the lamp to within about
a third of the bottom. (See Fig.) The thin chloroform-balsam is now to
be filtered into this lamp, very fine filtering-paper—through which a
little chloroform has first been passed—being used for the purpose. When
the lamp is full it must (deprived of its cap) be put in a warm place
until sufficient of the chloroform has evaporated to leave behind it a
fluid of the consistence of thin syrup.
Footnote 14:
Although we cannot too strongly insist upon the use of
chloroform-balsam wherever practicable, yet it sometimes happens in
the mounting of substances of _considerable thickness_, that after all
the chloroform has evaporated an insufficient amount of balsam is left
behind to fill up the cavity between slide and cover. In such cases,
therefore, it is advisable to use pure balsam, which may be done in
the following manner. The object having been previously thoroughly
dehydrated by immersion in absolute alcohol, is to be thence
transferred to a little good turpentine, or benzole, where it should
remain until perfectly transparent. It is now to be placed in the
centre of a slide which has been gently warmed, and a drop or two of
_fresh_ fluid balsam added, the greatest care being taken to prevent
the formation of air-bubbles. Should such arise they must be touched
with the point of a heated needle, which will cause them to burst and
disappear. The chief difficulty of the process has yet to be
encountered in the application of the cover; for it is during this
procedure that the development of air-bubbles is most likely to take
place. This annoyance may, however, be entirely avoided by taking the
simple precaution of dipping the cover into turpentine before it is
applied (§ 16), when it will be found that “you can’t get air-bubbles,
even if you try.” The courtesy of Mr. J. A. Kay, of Chatham, enables
us to give our readers the benefit of this practical “_wrinkle_.”
[Illustration: SPIRIT LAMP ADAPTED TO CONTAIN BALSAM.]
23. _Mounting in Balsam._—Let us now return to our sections which, it
will be remembered, were left in methylated spirit (§ 19). These we will
now mount in balsam, and although, of course, any number may be
proceeded with at the same time, yet to avoid confusion, in the
following directions one section only will be spoken of. This section
then is, with the perforated spoon, to be transferred to the bottle of
absolute alcohol, where it may remain for about an hour—considerably
less time is _actually_ required, but as from constant use the spirit
becomes weakened, it is as well to be on the safe side. It must now be
removed to the centre of a clean glass slip, and here the _plain_ end of
the spoon comes into use. If this be employed for effecting the
transfer, it will be found that when the section is being removed from
the alcohol it will bring along with it a small pool of the spirit. A
slight touch of the needle applied to the edge of the section will cause
it to float from the spoon on to the slide, at the same time carrying
the pool of alcohol with it, in which it will gently spread itself out
upon the slide without the faintest risk of injury. The superfluous
spirit is now to be drained off, and just as the section is becoming
glazed and sodden-looking (_not dry_) we must, by means of the long
glass stopper (§ 21), apply to it a large drop of clove oil. The oil,
however, should not be placed on the section, but be allowed to drop on
to the slide near to its margin. By gently tilting the slide the oil
will gradually insinuate itself _beneath_ the section and slowly ascend
through it to the surface. The slide should now be covered with a
bell-glass (or wine-glass), and about two minutes allowed for the oil
thoroughly to saturate the section. As much as possible of the
superfluous oil must then be drained off, and the remainder removed with
blotting paper. By means of the glass rod a small quantity of
chloroform-balsam is now taken from the spirit lamp which contains it,
and allowed gently to fall upon the section, which must then be covered
with a thin glass circle in the manner previously described (§ 16). When
the object is very fragile, it is a good plan, after draining off the
clove oil, to apply the cover directly upon the section, and then to
place a drop of the balsam near to the edge of the cover. This, by
capillary attraction, will speedily diffuse itself beneath the cover,
flowing over and surrounding the object, without in the slightest degree
disturbing its position. If, during the process of mounting, any
air-bubbles arise, we may view their development with equanimity, being
well assured that as the chloroform evaporates they too will quickly
disappear. When the mounting is completed, the slide should be roughly
labelled and placed on a warm mantel-piece for a few days to dry.
24. _Finishing the Slide._—In the course of two or three days it will be
advisable to take an old penknife, and after heating the blade in the
flame of a spirit lamp, gently to run the point of it round the margin
of the cover, so as to remove any excess of balsam which may have oozed
from beneath it. In a few days more, any remaining balsam may be
carefully scraped away with a cold knife. All remaining traces of balsam
are then to be removed from around the cover by means of a rag _just
moistened_ with methylated spirit, or, what is better, with a mixture of
equal parts of spirit and æther, after which the slide is to be
thoroughly washed in cold water. The slide is now in reality finished,
but, in order to give it a smart appearance, it is usual, with the
assistance of the turn-table, to run a ring of colored varnish round the
covering glass. A very useful varnish for the purpose is the _white zinc
cement_. To prepare this, dissolve an ounce of gum dammar in an ounce of
turpentine by the aid of heat. Take one dram of oxide of zinc and an
equal quantity of turpentine; rub them up together in a mortar, adding
the turpentine drop by drop, so as to form a creamy mixture perfectly
free from lumps or grit. One fluid ounce of the dammar solution
previously made must now gradually be added, the mixture being kept
constantly stirred (_Frey_). The cement, when made, should be strained
through a piece of fine muslin, previously wetted with turpentine, into
a small wide-necked bottle, which, instead of having a cork or stopper,
should be covered with a loose metal cap. Instead of a bottle, the
varnish may be kept in one of the collapsible tubes used by artists; but
though this plan is highly recommended by many, it is not without its
disadvantages. If the varnish becomes thick by keeping, a few drops of
turpentine or benzole well stirred in will soon reduce it to a suitable
consistence for use.
------------------------------------------------------------------------
PART II.
25. _Special Methods._—Having in the preceding pages entered at some
length into the general subject of section-cutting, it remains for us
now to consider those special methods of preparation which the
peculiarities of certain objects demand. In order to keep the bulk (and
consequent price) of this manualette within due bounds, we shall,
without further preface, proceed to the description of these methods, in
doing which every endeavour will be made to employ such brevity of
expression as may be consistent with perfect clearness of meaning. As
the most convenient plan, the objects here treated of will be arranged
in alphabetical succession.
26. _Bone._—Both transverse and longitudinal sections should be
prepared, the former being the prettier and most interesting. After
prolonged maceration in water, all fat, etc., must be removed and the
bone dried, when as thin a slice as possible is to be cut off in the
desired direction, by means of a very fine saw. If the section so
obtained be placed upon a piece of smooth cork it may, with the aid of a
fine file and the exercise of care, be further reduced in thickness. It
is then to be laid upon a hone moistened with water, and being pressed
gently and _evenly_ down upon it with the tip of the finger (protected,
if necessary, by a bit of cork or gutta-percha), it must be rubbed upon
the stone until the desired degree of thinness has been attained.
Finally, in order to remove scratches and to polish the section, it
should be rubbed upon a dry hone of very fine texture, or upon a strop
charged with putty-powder. After careful washing in several waters the
section must be allowed thoroughly to dry, when it may be mounted by the
_dry method_ in the following manner:—A ring of gold-size must, by means
of the turn-table, be drawn in the centre of a slide, and the slide put
away in a warm place for several days (the longer the better), in order
that the ring may become perfectly dry and hard. When this has been
accomplished the section is to be placed in the centre of the ring, and
a covering circle of the requisite size having been cleaned, this must
have a _thin_ ring of gold-size applied round its margin. The cover is
now to be placed in position and gently pressed down, a spring clip
being employed, if necessary, to prevent it from moving. In about
twenty-four hours another layer of the varnish should be applied, and
the slide afterwards finished in the manner already described (§ 24).
The above method is also applicable to the preparation of sections of
_teeth_ and also of _fruit-stones_ and other hard bodies, which are
incapable of being rendered soft enough for cutting.
As the process just described, however, is both troublesome and tedious,
it is much better for ordinary purposes to have recourse to the
_decalcifying method_, by which means sections in every way suitable for
the examination of the essential structure of bone may be obtained with
ease. To carry out this plan a piece of fresh bone should be cut into
small pieces and placed in a solution made by dissolving 15 grains of
pure chromic acid in 7 ounces of distilled water, to which 30 minims of
nitric acid s. g. 1.420 are afterwards to be added. Here they should
remain for three or four weeks, or until the bone has become
sufficiently soft to cut easily, the fluid being repeatedly changed
during the process. From this solution they must be transferred to
methylated spirit for a few days, when a piece may be selected, imbedded
in paraffine, and cut in the microtome (§ 12). Some of the sections
should be mounted, unstained, in spirit. For this purpose a cell of
gold-size, as above described, must first be prepared and filled _full_
of a mixture of spirit of wine one part, and distilled water three
parts. Into this the section must be carefully placed and the cover
applied, the same precautions for the exclusion of air-bubbles being
taken which were recommended when speaking of mounting in glycerine (§
16). When the cover is in position a ring of gold-size must be laid on,
repeated when dry, and the slide afterwards finished in the ordinary
manner. It will also be advisable to stain some of the sections with
carmine (§ 14), or picro-carmine (§ 42), and mount them in glycerine.
_Teeth_ may also be treated by the decalcifying method, but in this case
it must be remembered that the enamel will dissolve away.
27. _Brain._—The best hardening fluid is that recommended by Rutherford,
and is made by dissolving 15 grains of pure chromic acid and 31 grains
of crystalized bichromate of potash in 43 ounces of distilled water.
Small pieces of brain, which have previously been immersed for
twenty-four hours in rectified spirit, should be placed in about a pint
of this solution, where they must remain for five or six weeks, the
fluid being repeatedly changed during the process. If by this time they
are not sufficiently hard the induration must be completed in alcohol.
Sections are easily cut in the microtome by the paraffine method (§ 12).
These may advantageously be stained in a solution of aniline blue, made
by dissolving 1-1/2 grain of aniline blue in 10 ounces of distilled
water, and adding 1 drachm of rectified spirit (_Frey_). As this stain
acts very rapidly two or three minutes’ immersion will generally be
found long enough. The sections must then be mounted in balsam (§ 23).
28. _Cartilage._—The method to be employed in the preparation of
cartilage will entirely depend upon the nature of the staining agent, to
the action of which the sections are to be submitted. Thus, if the
elegant _gold method_ is to be followed, it is necessary that the
cartilage should be perfectly fresh; whilst if any of the other staining
agents are to be employed the tissue may have been previously preserved
in alcohol. An excellent object on which to demonstrate the gold process
is to be found in the articular cartilage of bone. It is a very easy
matter to obtain from the butchers the foot of a sheep which has just
been killed. The joint is to be opened, and the bones dissociated, when
they will be seen to have their extremities coated with a white
glistening membrane—this is the _articular cartilage_. Exceedingly thin
slices must be at once cut from it, and as only small sections are
required, a sharp razor may be used for the purpose, the blade being
either dry or simply wetted with distilled water. The sections as cut
are to be transferred to a small quantity of a half per cent. solution
of chloride of gold in a watch glass. Chloride of gold may be purchased
in small glass tubes hermetically sealed, each tube containing 15
grains, and costing about 2s. If, however, the student requires only a
small quantity of the staining fluid he need not be even at this small
expense, for as photographers for the requirements of their art always
keep on hand a standard solution of chloride of gold of the strength of
one per cent., a little of this may readily be obtained, and diluted to
the required degree. After the sections have been exposed to the action
of the staining fluid for about ten minutes they may be transferred to a
small beaker of distilled water, and exposed to diffused light for about
twenty-four hours, when they must be mounted in glycerine (§ 16).
Sections of cartilage may also be examined, without being stained, in
which case the field of the microscope should be only very feebly
illuminated. Or carmine staining (§ 14) may be resorted to—these
sections show well in glycerine, or if the staining be made very deep,
even Canada balsam may be employed, and with fair results.
Microscopists are indebted to Dr. Frances Elizabeth Hoggan for the
description of a new method of staining, which we have found especially
suited to the treatment of cartilage. The agent employed is _iron_, and
the process, which is very simple, is as follows. Two fluids are
necessary—(1) tincture of steel; (2) a two per cent. solution of
pyrogallic acid in alcohol. A little of the former is to be poured into
a watch glass, and into this the sections, after having been previously
steeped in alcohol for a few minutes, are to be placed. In about two
minutes the iron solution is to be poured away and replaced by solution
No. 2. In the course of a minute or two the desired depth of colour will
have been produced, when the sections are to be removed, washed in
distilled water, and mounted in glycerine. The results obtained by this
process are very beautiful, the colour produced being a very fine
neutral tint, of delightful softness. The process also answers admirably
in the case of morbid tissues, and we have now in our possession some
sections of ulcerated cartilage tinged by the iron method, in which the
minute changes resulting from the ulcerative disintegration are brought
out with wonderful distinctness.
As the structure of cartilage differs according to its purpose and
situation, the student will find his time profitably employed in a
careful examination of the following forms (α) _hyaline_—articular and
costal; (β) yellow fibro-cartilage—epiglottis, or external ear; (γ)
_cellular_—ear of mouse. Sections of the _intervertebral_ ligaments
should also be made, in which the different kinds of cartilage may be
examined side by side with each other.
29. _Coffee Berry_ affords sections of great beauty. The _unroasted_
berry should be soaked for hours or days in cold water until
sufficiently soft; then imbedded in paraffine, and cut in the microtome
(§ 12), the section being made in the direction of the long axis of the
berry. Put up in glycerine, or stain rather strongly with carmine, and
mount in balsam. The same method of treatment may also be applied to
other hard berries or _seeds_.
30. _Fat._—Adipose tissue may be hardened in alcohol, cut in paraffine,
and mounted in glycerine. If the tissue has been injected the sections
may be mounted in balsam, and are then very beautiful objects, showing
the capillary network encircling the fat cells.
31. _Hair._—Longitudinal sections are readily made by splitting the hair
with a sharp razor. It is more difficult to cut the hair transversely.
This, however, may easily be done in the following manner. The hairs
having previously been well soaked in æther to remove all fatty matters,
a sufficient number of them must be selected to form a bundle about the
thickness of a crow quill. This bundle, after being tied at each
extremity with a bit of thread, is to be immersed for several hours in
strong gum (§ 18,) to which a few drops of glycerine have been added. On
removal, the bundle must be suspended by means of a thread attached to
one end of it, in a warm place until sufficiently hard, when it is to be
imbedded and cut in paraffine (§ 12). Each section, as cut, is to be
floated off the knife into methylated spirit. From this it is with the
aid of the spoon (§ 14) to be transferred to a slide, the spirit tilted
off, a drop of absolute alcohol added, when, after a minute or two, this
also is to be drained off, the section treated with clove oil, and the
mounting completed as described in § 23.
32. _Horn_ varies very much in consistence, in some instances having a
cartilaginous character, whilst in others it is almost bony. In the
latter case, sections will have to be ground down in the manner
explained when speaking of bone (§ 26). Where the texture is less dense,
recourse may be had to prolonged steeping in hot or boiling water; in
some cases it will be necessary to continue the immersion for several
hours. When sufficiently soft the piece of horn may, by means of bits of
soft wood, be firmly wedged into the tube of the microtome, and sections
cut with a razor, or what is better, with a broad and very sharp chisel.
The sections are to be put between glass slips, held together by
American clips (or pegs), and put away for two or three days in order to
become thoroughly dry. After well soaking in good turpentine or benzole,
they must be transferred to slides, the superfluous turpentine drained
off, and chloroform-balsam added, etc. (§ 23). Sections of horn should,
of course, be cut in different directions, but for examination with the
polariscope those cut transversely yield by far the most magnificent
results. _Hoofs_, _whalebone_, and allied structures should also be
treated by the above method.
33. _Intestine._—The method to be pursued with _sections_ has already
been described (§ 18). The ileum, however, is a very pretty object when
a portion of it is so mounted as to show the _villi erect_. To do this
it is necessary to cement to the slide, by marine glue, a glass cell of
sufficient depth. This should have been prepared some time beforehand,
so that the cement may be perfectly dry and hard. The cell is now to be
filled with turpentine, and the piece of ileum (having been previously
passed through methylated spirit and absolute alcohol into turpentine)
is gently placed into it, having the villi uppermost; pour some pure and
rather fluid balsam on the object at one end, and gradually incline the
slide, so as to allow the turpentine to flow out at the opposite side of
the cell, till it is full of balsam. Then take a clean cover, and having
placed upon it a small streak of balsam from one end to the other, allow
it gradually to fall upon the cell, so as to avoid the formation of
air-bubbles (§ 17), and finish the slide in the usual manner.[15] Or,
the intestine may be dried, and mounted _dry_, in a cell with a
blackened bottom, for examination as an opaque object.
Footnote 15:
Ralf.
34. _Liver._—Small pieces of liver may be very successfully hardened by
immersion in alcohol, beginning with weak spirit and ending with
absolute alcohol. Cut and mount as usual.
35. _Lung_ must be prepared in chromic acid (§ 5). For the cutting of
sections the freezing microtome (§ 18) is of especial value, and should,
therefore, be used. If, however, the student be not provided with this
instrument, he must proceed as follows. A small piece of lung,
previously deprived of all spirit, is to be immersed until thoroughly
saturated in solution of gum (§ 18). A small mould of bibulous paper (§
2), only just large enough to receive the piece of tissue, having been
prepared and filled with the mucilage, the specimen is to be transferred
to it. The mould, with its contents, is now to be placed in a saucer,
into which a mixture of about 6 parts of methylated spirit and 1 part of
water (_Schäfer_) is to be poured until the fluid reaches to within
about a third of the top of the paper mould. In the course of several
hours the surface of the mucilage will begin to whiten and solidify. As
soon as this occurs more dilute spirit must be poured into the saucer,
until the mould is completely submerged. In a day or two the gum will be
found to have acquired a suitable consistence for cutting, when it must
be removed from the spirit, the paper mould peeled off, and the mass
imbedded and cut in paraffine, the sections being afterwards treated as
if they had been obtained by the freezing method (§ 18). If the
solidification of the gum should proceed too slowly, a few drops of pure
spirit may be added to the contents of the saucer. If, on the other
hand, the gum should become overhard, it will be necessary to put into
the saucer a few drops of water, and repeat this until the required
consistence be obtained.
36. _Muscle._—Harden in chromic acid, and cut in paraffine. Transverse
sections may be made to show the shape of the fibrils. Longitudinal
sections will only be required in the case of injected tissues, when
such sections will be found very elegant, showing, as they do, the
elongated meshes of capillaries running between and around the muscular
fasciculi. Mount in glycerine or balsam. To see the transverse striæ
characteristic of voluntary muscle, a very good plan is to take a bit of
pork (cooked or fresh), and by means of needles to teaze it out into the
finest possible shreds. If these be examined in water or glycerine, the
markings will be shown very perfectly.
37. _Orange-peel_, common object though it be, is not to be despised by
the microscopist. Transverse sections must be prepared by the gum method
(§ 35). These sections are not to be subjected to the action of alcohol
(as this would destroy the colour), but after _drying_ between glass
slides they must be soaked in turpentine and mounted in balsam. We shall
then have a good view of the large globular glands whose office it is to
secrete that essential oil upon which the odor of the orange depends.
38. _Ovary_ may be prepared in the same manner as liver (§ 34).
Sections, which are to be cut in paraffine, may be stained with carmine,
and mounted in glycerine or balsam. Apart from all scientific value, we
know of no slide for the microscope which, even as a mere object of
show, surpasses in beauty a well-prepared section of _injected_ ovary,
showing the wondrous Graafian vesicles, surrounded by their meandering
capillaries.
39. _Porcupine Quill._—Soften in hot water, cut in paraffine, and mount
in balsam. Much (in our opinion _too_ much) lauded as an object for the
polariscope.
40. _Potato._—From the large amount of water which it contains thin
sections cannot be cut from the potato in its natural state. It must,
therefore, be partially desiccated, either by immersion in methylated
spirit for a few days or by exposure to the air. Sections may then
readily be obtained by imbedding and cutting in paraffine. Such sections
mounted in balsam are very beautiful, the starch being seen _in sitú_,
whilst if polarized light be employed each granule gives out its
characteristic black cross.
41. _Rush_ is to be prepared and cut as orange-peel (§ 37). Transverse
sections of this “weed” furnish slides of the most exquisite beauty.
42. _Skin._—To prepare skin for section a piece is to be selected which,
after having been boiled for a few seconds in vinegar, must be stretched
out on a bit of flat wood, and being maintained in position by pins be
allowed to remain until thoroughly dry. Then imbed in paraffine, and cut
_exceedingly_ thin transverse sections. These may be stained in carmine,
but more beautiful results are obtained if picro-carmine be employed.
Sections of skin, when stained by this agent are much increased both in
beauty and instructiveness; for the several constituents of the tissue
becoming tinged with different colours are readily distinguishable from
each other, whilst the contrast of colouring forms a pleasing picture to
the eye. The method of preparing picro-carmine is very simple, though it
sometimes yields a solution not altogether satisfactory. The best
formula with which we are acquainted is that given by Rutherford,[16]
and if due care be taken in following it out failure will generally be
avoided. “Take 100 c.c. of a saturated solution of picric acid. Prepare
an ammoniacal solution of carmine, by dissolving 1 gramme in a few c.c.
water, with the aid of excess of ammonia and heat. Boil the picric acid
solution on a sand bath, and when boiling add the carmine solution.
Evaporate the mixture to dryness. Dissolve the residue in 100 c.c.
water, and filter. A clear solution ought to be obtained; if not, add
some more ammonia, evaporate, and dissolve as before.” Sections may be
exposed to the action of this fluid for a period varying from fifteen to
thirty minutes, then rapidly washed in water, and mounted in glycerine.
They may also be mounted in balsam, care being taken in that case to
shorten as much as possible the period of their immersion in alcohol, so
that no risk may be run of the picric acid stain being dissolved out.
Footnote 16:
“Practical Histology,” 2d edit. p. 173.
If it is intended to study the structure of the skin with anything like
thoroughness, portions must of course be examined from different
localities, in order that its several varieties and peculiarities may be
observed. Thus the _sudoriforous_, or sweat glands, may be found in the
sole of the foot, whilst the _sebaceous_ glands are to be sought in the
skin of the nose. The _papillæ_ are well represented at the tips of the
fingers,[17] whilst the structure of the shaft of the _hair_, together
with that of the follicle within which its root is enclosed, as also the
muscles by which it is moved, are to be studied in sections of skin from
the scalp or other suitable locality.
Footnote 17:
It is well, in connection with these papillæ, to bear in mind a fact
pointed out by _Frey_, namely, that the tips of the fingers frequently
become, _post-mortem_, the seat of extensive natural injections;
hence, in sections from this region, we frequently obtain good views
of distended capillaries without having been at the trouble of
previously injecting them.—_Frey_, “Microscopical Technology.”
43. _Spinal Cord._—The spinal cord, say of a cat or a dog (or if
procurable, of man), after being cut into pieces about half an inch in
length, may be hardened in the usual chromic acid fluid (§ 5). As it is
peculiarly liable to overharden and become uselessly brittle, the
process must be carefully watched. Its further treatment is the same as
that of brain. These sections may be stained very satisfactorily by the
_ink process_, for communicating details of which we are indebted to the
kindness of Dr. Paul, of Liverpool. The agent usually employed is
Stephenson’s blue-black ink, which, for this purpose, must be quite
fresh. As in the case of carmine, two methods of staining may be
adopted—either rapid, by using concentrated solutions, or more
prolonged, according to degree of dilution. For the reasons previously
given (§ 14), slow methods of staining are always to be preferred, as
yielding the most beautiful results, yet, for the purposes of
preliminary investigation, it is often convenient to have recourse to
the quick process. To carry out the latter plan, an ink solution of the
strength 1 in 5—10 parts of water is to be freshly prepared, and the
sections exposed to its action for a few minutes. For gradual staining
the dilution must be carried to 1 in 30—50, and the time of immersion
prolonged to several hours, the sections being occasionally examined
during the staining, so that they may be removed just as they have
acquired the desired tint. When a satisfactory coloration has been
obtained, the preparations should be mounted in dammar or balsam (§ 23).
One advantage of this method of staining is, that definition is almost
as good by artificial light as by day.
44. _Sponge_ may readily be cut after being tightly compressed between
two bits of cork; or its interstices may be filled up by immersion
either in melted paraffine (§ 11) or in strong gum (§ 18), and then cut
as usual.
45. _Stomach_ requires no special method of hardening (chromic acid).
Sections should always, when practicable, be cut in the freezing
microtome. In default of this, proceed in the manner as directed for
lung (§ 35). Both vertical and horizontal sections will, of course, be
required. If the preparation has been injected, the latter are
particularly beautiful. Stain with carmine or aniline blue (§ 27), and
mount—if for very close study, in glycerine—if injected and for a “show”
slide, use balsam.
46. _Tongue._—Harden in chromic acid, imbed and cut transverse sections
in paraffine. As, however, the paraffine is apt to get entangled amongst
the _papillæ_, whence it is afterwards with difficulty dislodged, it
will be as well before imbedding to soak the tongue in strong gum for a
_few minutes_, and afterwards immerse in alcohol till the gum becomes
hardened, so that the delicate papillæ may thus be protected from the
paraffine by a surface-coating of gum. The best staining agent is
picro-carmine (§ 42). Sections of _cat’s_ tongue near the root, when
thus stained, furnish splendid objects. Sections should also be made of
the _taste-bulbs_, found on the tongues of rabbits. These are small oval
prominences, situated one on each side of the upper surface of the
tongue near its root. They should be snipped off with scissors, and
vertical sections made in the direction of their long axis. Stain with
carmine or picro-carmine, and mount in glycerine or balsam.
47. _Vegetable Ivory_.—After prolonged soaking in cold water may readily
be cut in the microtome. The sections should be mounted in balsam, and
though not usually regarded as polariscopic objects, nevertheless, when
examined with the _selenite_, yield very good colours.
48. _Wood._—Shavings of extreme thinness may be cut from large pieces or
blocks of timber, by means of a very sharp plane. In this way very good
sections may be procured of most of the common woods, as oak, mahogany,
“glandular wood” of pine, etc. Where however, the material to be
operated upon takes the form of stems, roots, etc., of no great
thickness, they should, after having been reduced to a suitable
consistence (§ 4), be imbedded in paraffine, and cut in the microtome.
Before imbedding it must not be forgotten to immerse the wood to be cut
in weak gum-water (§ 11), this precaution being of great importance,
especially in the case of stems, etc., the bark of which is at all rough
and sinuous. If the sections are to be mounted _unstained_, they are
usually put up in weak spirit (§ 26). A very general method also of
dealing with this class of objects is to mount them _dry_ (§ 26). This
plan, however, cannot be recommended, for however thin the sections may
be, the outlines, when this process is adopted, always present a
disagreeable black or blurred appearance. To avoid this we may have
recourse to Canada balsam, but the ordinary method of employing it must
be slightly modified, a drop of chloroform being substituted for the
clove oil (§ 23), otherwise this latter agent will cause the section to
become so transparent as to render minute details of structure difficult
to recognize. A better plan, perhaps, is to stain the section with
carmine or logwood, and mount in balsam by the ordinary process. The
best course to follow, however, especially in the case of transverse
sections, is the _double staining_ method.[18] For this purpose the
sections in the first place must be subjected to the action of a
solution of chloride of lime (1/4 oz. to a pint of water) until they
become thoroughly bleached. They must then be soaked in a solution of
hyposulphite of soda (one drachm to four ounces of water) for an hour,
and after being washed for some hours, in several changes of water, are
to be transferred for a short time to methylated spirit. Some red
staining fluid is now to be prepared by dissolving half a grain of
Magenta crystals in one ounce of methylated spirit. A little of this
solution being poured into a small vessel of white porcelain (§ 14), the
sections are to be immersed in the dye for about thirty minutes. They
are now to be removed, and after _rapid_ rinsing in methylated spirit to
remove all superfluous colour, they must be placed in a blue staining
fluid made by dissolving half a grain of aniline blue in one drachm of
distilled water, adding ten minims of dilute nitric acid and afterwards
sufficient methylated spirit to make two fluid ounces. The sections must
be permitted to remain in this solution for a very short time only, one
to three minutes being generally sufficient, for as the action of the
dye is very energetic, it will, if too long exposure be allowed,
completely obliterate the previous coloration by the magenta. After
being again _rapidly_ rinsed in methylated spirit, as much of this as
possible must be drained off, and the sections put into oil of
_cajeput_, whence, in an hour, they may be transferred to spirits of
turpentine, and after a short soaking, mounted in balsam.
Footnote 18:
See a paper by Mr. Styles in the “Pharmaceutical Journal,” also
“Monthly Microscopical Journal” for August, 1875. [For a very
exhaustive paper on this subject by the late Dr. Beatty, of Baltimore,
Md., see “American Journal of Microscopy” for June, 1876.
If the student will carefully carry out the above process, his trouble
will be amply repaid by the beautiful results obtained, for by its means
he may, with ease, prepare for himself a series of slides of such value
as to constitute a worthy addition to his cabinet.
The preceding list by no means represents all the objects, sections of
which will be found interesting to the microscopic student. Such was not
its purpose—had it been so, the enumeration might have been prolonged
almost indefinitely. The end in view was to bring under the notice of
the reader only those substances the cutting of which is accompanied by
difficulty; and even of this class the space at our disposal has been so
limited that we have been unwillingly compelled to pass over many, and
dwell only on such as possess a typical character.
------------------------------------------------------------------------
NOTES.
----------
NOTE A., Page 14.
RANGOON oil is not found in this market. It is a variety of naphtha or
petroleum found at Rangoon, in Asia, and contains a variable percentage
of solid paraffin. Kerosene with a very little paraffin dissolved in it,
would undoubtedly answer the same purpose.
Good sperm oil or tallow (free from salt) will serve quite as well. It
is scarcely necessary to caution the reader that all oily matter should
be removed from the knife before it is used for cutting sections. This
is best effected by dipping it in hot water and carefully wiping it.
NOTE B., Page 15.
METHYLATED spirit is alcohol which contains a small percentage of
methyl. This renders it unfit for use in the manufacture of liquors or
even chloroform, but does not injure it as a preservative fluid. The
scientific men of Great Britain have the privilege of obtaining this
fluid free from duty, and consequently they make very general use of it.
In this country the student of science has no such privilege, and must
use the more expensive alcohol. This however is but of a piece with
other restrictions placed upon scientific pursuits in this country,
where men like Marshall Jewell and Hannibal Hamlin have influence enough
to pass laws which prohibit the exchange by mail of microscopic
specimens, which pass freely in every other civilized country.
True methyl alcohol, or methylated spirits, is not to be had in this
country, and therefore whenever this liquid is directed to be used in
English books the American microscopist must substitute good alcohol.
What is sold for “methyl alcohol” is wood spirit or wood naphtha, and
will not serve the purposes of the microscopist, though it answers well
enough for spirit lamps, making varnishes, etc.
NOTE C., Page 20.
THE method described in the text is generally known as the “free-hand”
method, and although it is not generally employed for the production of
cabinet specimens or slides that are intended for sale, yet it possesses
so many advantages that every student should endeavor to acquire the art
of cutting sections without the use of the microtome. For purposes of
study such sections are quite equal to the best of those usually found
on sale, and for the encouragement of beginners we may add that we have
seen sections cut by the free-hand method which for size, thinness and
accuracy, excelled any that we have ever seen produced with the
microtome. Indeed Schäffer distinctly affirms that no microtome can
equal the skilled hand in the production of thin sections.
NOTE D., Page 22.
THE mere existence of a binding screw should be no objection, and where
bushings are employed it is sometimes necessary. But as the author well
observes, a binding screw cannot be used to secure histological
preparations.
NOTE E., Page 22.
A STILL better system is where the section-knife is held in a frame
which slides on the plate so that the edge is kept just above the
surface. In this way the edge of the knife is never dulled by grinding
on glass or metal. A common mistake in this form, however, is to raise
the cutting edge too far above the surface of the plate. This gives rise
to great irregularities.
NOTE F., Page 22.
AS the size of the sections which it may be desired to cut, varies
within wide limits, the best microtomes made in this country are
furnished with as many as three tubes or “bushings,” fitting one within
the other, and in this way the hole may be partially filled up and thus
reduced in size. This enables us to obtain a range of from one inch and
a quarter, suitable for large histological preparations, to one-eighth
of an inch for such objects as hair sections.
NOTE G., Page 37.
KNIVES FOR CUTTING SECTIONS.
SUCCESS in cutting sections depends more upon the excellence of the
knife that is used than upon any other one point, and therefore the art
of keeping the knife in good order is one which should be acquired by
every microscopist, even at the cost of considerable time, study and
labor. As a stepping stone to the acquisition of this art, there are
certain general principles which must be thoroughly understood before we
can hope for success in practice. These principles are simple, and when
once clearly stated, quite obvious, but they are frequently overlooked.
Cutting instruments act in two ways—either as simple wedges or as a
series of wedges, the latter being generally known as the saw-like
action. It is frequently stated in school philosophies that the edges of
_all_ cutting instruments are toothed like a saw, and that they act like
a saw; this is evidently not the case with many, such as the axe, the
chisel, the plane, etc., and it is not even true of several to which it
is frequently applied, such as the razor, the penknife, etc. The action
can be said to be “saw-like” only when the teeth act as independent
wedges and remove a small portion of the substance acted upon, the
portion being known as _saw-dust_. Now in cutting sections of wood with
a sharp razor, where but a single stroke is used to carry the cutting
edge across a section, say half an inch in diameter, the edge of the
razor does not act like a saw; none of the material is removed except
the section itself; in other words, there is no saw-dust. That the edge
of the knife or razor is not even, is very easily proved by means of the
microscope, but it will be found that the notches and projections form a
series of lancet-shaped knifelets, each one of which acts just as does
the blade of a penknife when drawn over a piece of wood or a quire of
paper. Now if we lay a quire of paper on a board and draw the edge of a
penknife over it, we will cut down through the paper, not by sawing but
by a moving wedge. It is precisely in this way that the little
lancet-shaped teeth on the edge of a razor act when the latter is used
to cut a section.
If we further examine the edge of the razor, we will find that the
bottoms of the notches are comparatively blunt; if therefore we press
the razor directly forward we soon press these blunt portions of the
general edge against the material to be cut, and the resistance becomes
so great that the material is actually crushed or torn. This would not
happen if we had an absolutely perfect edge—one without any notches.
This being practically unattainable, however, we must endeavor to make
up for it by a sawing motion, (though not a sawing action) at the same
time seeking to avoid as much as possible those saw-like features which
render this motion necessary. We have dwelt at length upon this point
because so much has been written comparing the edge of a razor to that
of a saw, that some persons actually advise us to give the edges of our
knives and razors a saw-like character, under the impression that by so
doing they will work better. This is a fallacy; the best work will be
obtained from the smoothest and most perfect edge, provided the latter
is thin enough.
If the material of which our cutting instruments are made would allow
it, that is to say if it was perfectly hard, perfectly rigid and
incapable of being crushed or torn apart, the proper shape of a cutting
instrument would be a wedge of the most acute form possible. But since
the steel which we use is limited in its hardness and its power of
resisting crushing and bending influences, the blades of our knives must
have a certain strength or thickness, depending upon the character of
the material to be cut. For very hard substances the knife must be stout
and the edge ground to a comparatively obtuse angle; for moderately soft
substances, such as wood, the angle may be more acute, while for very
soft tissues the thinnest blade and the sharpest edge are most suitable.
[Illustration: Fig. 1.]
The various angles to which cutting tools of different kinds are ground
are shown in the accompanying illustrations, which explain themselves.
[Illustration: Fig. 2.]
In the diagrams just given, the sides of the wedge are carried in
straight lines to the very edge. In practice, however, this is never
done, there being two strong objections to such a form. In the first
place if the sides were as shown in the figures, every time the tool was
ground or set, the entire side would have to be ground off, and this
would involve great labor. But another difficulty, quite as great, would
occur in the use of the instrument, for since the wedge keeps growing
constantly and proportionately thicker, the friction between the sides
of the wedge and the material would rapidly become very great. Therefore
most cutting edges are ground to a second angle, much more obtuse than
the first, and it is this second angle which forms the true cutting
edge. And it is obvious that the mass of material required to carry out
the ultimate angle in any case is wholly unnecessary, for if the tool be
strong enough at _a b_ (Fig. 2) it will certainly be strong enough at _c
d_. An extreme illustration of this principle is shown in the cold
chisel used for cutting iron, the edge of which is shown in Fig. 2, and
the angle of 80° to which it is ground is carried out by dotted lines.
The angle which the sides of the chisel (not the sides of the extreme
cutting edge however) make with each other is 25°.
[Illustration: Fig. 3.]
[Illustration: Fig. 4.]
There are two methods by which this change of angle may be made, one of
which is shown in the cold chisel just figured, and the other in the
razor of which a section is shown in figures 3 and 4. In the case of the
cold chisel it will be seen that the tool is first formed to a thin
straight wedge which is afterwards changed to one that is much more
blunt. This answers very well where the ultimate angle is comparatively
large or blunt, as is the case with penknives, table-knives,
carving-knives, etc. But where the ultimate angle is very small this
plan does not answer well, and the method shown in Fig. 3 is generally
adopted. Here the ultimate angle is such that lines touching the extreme
edge and the back of the blade are perfectly straight and form the
actual cutting angle. In this case, therefore, the relief is obtained by
hollowing out the sides of the blade, and this is done to various
extents, the extremes being shown in figures 3 and 4. Fig. 3 shows a
section of a razor ground on a stone 12 inches in diameter, which is as
large as is generally used for this purpose. Fig. 4 is a section of a
razor ground on a four-inch stone—the smallest in general use. This
method of changing the angle is of course substantially the same as the
first, merely differing in the mechanical device used, but it affords
this important advantage that in the subsequent honing and stropping
processes the back of the razor forms a perfect guide by which the
ultimate angle may be determined. This is not the case with the cold
chisel or the carving-knife, in both of which cases the eye and hand
alone determine the cutting angle, which is therefore apt to become
irregular or even rounded—the worst form of all. This will be more
easily understood from the following engravings where A, Fig. 5, shows a
penknife blade, as applied to an oilstone for the purpose of giving it
the final edge. The angle here shown is considerably greater than that
generally used, but the principle is the same. It is evident that if, in
moving the blade back and forth on the stone we allow it to rock or
change the angle which the blade makes with the stone’s surface, the
edge will become rounded as shown in Fig. 6—a form which for delicate
work is useless.
If the blade be laid down flat as at B, Fig. 5, the labor of wearing
away the superfluous metal will be enormous. But if the blade be hollow
as Fig. 7, (where the hollow is, for clearness of illustration, slightly
exaggerated) then the ultimate angle may be formed quickly and
accurately, there being no danger of the angles being changed on account
of want of skill on the part of the operator.
[Illustration: Fig. 5.]
[Illustration: Fig. 6.]
[Illustration: Fig. 7.]
[Illustration: Fig. 8.]
In some cases one side is left quite flat and the other side is ground
to two or sometimes three angles, as in the carpenter’s chisel, a
section of which is shown in Fig. 8, where _a a_ shows the angle made by
the body of the blade; _b b_ the angle of the bevel formed by the
grindstone, and _c c_ the angle of the cutting edge which is formed by
the oilstone. In this case the flat side A B forms a straight guide and
enables the workman, in cutting, to make straight, clean work. In
sharpening such a tool on the oilstone the flat side is always laid flat
on the oilstone and merely smoothed off,[19] and the skill of the
workman is chiefly shown in the accuracy with which he forms the angle
_c_ B _c_. The dexterity with which skilled workmen can effect this by
the hand alone is astonishing, but in the hands of those who have had
but little practice the edge infallibly becomes rounded as shown in Fig.
6. Therefore wherever a knife with a flat side is used, no attempt
should ever be made to grind this flat side on the hone. Such an attempt
will almost always result in an edge so irregular that it will be almost
impossible to cut a good section with it. But if the under side of the
knife be left truly flat, we will have the very best tool that can be
had for use with the microtome. Unfortunately, however, it is very
difficult to get a knife with a truly flat surface, that is, one that is
perfectly “out of wind” as mechanics say. As a general rule the flat
sides of knives are not true planes but irregularly curved surfaces;
consequently when laid on a plane surface, such as the table of a
microtome, they rock, and the edge is in contact with the table at some
points and separated from it at others, and the points which are in
contact change as the knife is moved diagonally across the table. This
makes the section irregular and worthless. But a really good knife with
a perfectly true flat side is such a valuable tool that a good deal of
effort may be profitably expended in getting it, and when once obtained
never let the flat side be touched by a grindstone, or more than touched
by a hone. It is not absolutely necessary, however, that the sides of a
delicate knife should be curved, provided the knife is used for cutting
very soft materials. An angle of 10° or even 5° gives a pretty stout
blade when carried back three-quarters of an inch, as may be seen by
examining Fig. 1, and the guiding action of the very thick back may be
obtained by means of an artificial guide, applied like the brass backs
of tenon saws. Such an arrangement is shown in Fig. 9, and has been
found very effectual.
Footnote 19:
Holtzapffel tell us that chisels that are required for paring across
the end grain of moderately soft wood are considered to hang better to
the work when they have a very slight keen burr or wire edge thrown up
on the face or flat side of the tool. But this does not apply to
section knives.
When the knife is to be honed, the back is applied and fastened by means
of two or three set screws. When laid on the hone, the edge of the knife
and the lower surface of the back form the guide and regulate the
cutting angle. In this way we can use a broad, thin blade and yet secure
great accuracy in honing it. Such a blade has this advantage also, that
it is not so liable to be rounded and thus injured by stropping as one
in which the relief is obtained by hollowing out the sides.
[Illustration: Fig. 9.]
Such, then, are the general principles which govern the construction of
cutting tools, including the section knives used by microscopists. We
will now give a few practical directions for selecting a knife and
putting and keeping it in order.
In selecting a knife for cutting sections, regard must be had to the
texture of the material that is to be cut. To attempt to cut delicate
sections of soft tissues with the stout knives which are suitable for
cutting sections of woody fibre would be to destroy the sections, while
to reverse the operation and cut sections of wood with thin, delicate
knives would result in the ruin of the knife. We have seen a most
excellent knife seriously injured by an attempt to cut material that was
too hard for it. The knife was very thin, and had proved most excellent
in cutting sections of such material as kidney, liver, etc. An attempt
was made, against our protest, to cut a section of an apple-shoot, the
wood of which was mature. Before the knife had gone half way across, it
bent, dug into the wood and broke, leaving an ugly gap in the middle.
Experienced section-cutters know this very well, but young microscopists
are not so familiar with the facts just stated, and the point is too
important to be overlooked. Those, therefore, who devote themselves to
microscopical studies, or who expect to make sections of materials of
several kinds, differing in hardness, etc., must provide themselves with
knives of different degrees of strength.
For common work, good razors are as good as anything, provided they can
be obtained with straight edges. Where razors are not suitable, recourse
must be had to the surgical instrument maker, though we are sorry to say
that there are but few in this country that know how to forge, temper
and grind a decent knife. Most of our dealers in instruments do not make
the instruments they sell; they import the goods they sell with their
names stamped on, and thus get a reputation as manufacturers; a special
order they are unable to fill respectably. There are some exceptions,
but of the majority of dealers what we have written above is true.
The points which specially demand attention in a knife for cutting
sections are these: 1. Quality of the steel used; 2. Temper; 3. Form of
the blade.
Of the quality of the material of which a knife is made, nothing can be
determined except by actual trial. The old tests of staining with acids,
examining with the microscope, etc., are worthless, or at least too
crude and uncertain to be of any practical value. Color changes with the
degree of polish that is produced, and, in short, there is no reliable
guide. The purchaser must depend entirely upon the reputation of the
manufacturer. There is plenty of good steel to be found; the trouble
lies with the cutlers. They are careless and in haste, and as a
consequence they burn the steel or fail to work it sufficiently, and the
result is a useless tool.
The steel may be of the very best quality, however, and well forged, and
yet the knife may fail from being badly tempered—too soft or too hard.
If too soft, the edge is soon dulled; the knife requires to be
frequently honed, and the time wasted in keeping it in order is a
serious drawback. If too hard, it is impossible to give it a keen edge,
for the metal crumbles away as soon as it is honed or stropped very
thin, and the edge becomes ragged and dull. Good steel, well forged, may
be so tempered that it will neither crumble nor become rapidly dull.
Much may be learned on this point from careful inspection of the edge,
and trial on a piece of horn, such as an old razor handle. When drawn
over a clean piece of horn once or twice the edge of a soft knife is
completely dulled; if well-tempered it should scarcely lose its
keenness. Again, when laid flat on the thumb nail and pressed, the edge
ought to bend up without breaking or crumbling, and at once, when the
pressure is removed, resume its original shape. The extent to which
extreme hardness and durability, or toughness, may be combined, is well
shown in the famous Toledo sword-blades. One of these blades will shear
through an iron nail without having its edge perceptibly dulled, and yet
so tough and springy is it that it may be coiled up into a hoop of
several folds without breaking or receiving any permanent set. If we
could only get section knives of stuff like this, it would be a pleasure
to work with them.
After all, however, the only test of these two points, material and
temper, is a fair trial in actual practice. Of the form of the blade,
however, it is easy to judge, and there are a few points which are
frequently overlooked and which give rise to errors that are attributed
to other causes.
In the text we are told that “It is essentially necessary that the back
and edge of the blade be strictly parallel to each other.” The author
undoubtedly knew what the correct form should be, but the definition he
has given is not a correct mathematical statement of the conditions
involved. These are as follows:
_The edge must form a straight line, and both the edge and the under
side of the back must lie in the same plane._
Let us consider these conditions somewhat carefully.
Since the hole in which the material to be cut is imbedded, is always
circular, and since the blade is generally pressed forcibly against the
table of the microtome, it is obvious that if the edge of the knife be
curved it will sink into the hole to a greater or less depth according
as it is further from the centre of the hole or nearer to it. To explain
this let us refer to the greatly exaggerated drawing given in figure 10
where the edge of the knife, _a_, _b_, is shown in two different
positions covering the hole of the microtome. The least degree of
reflection will show that as the knife passes towards the centre from _a
b_ to _aʹ bʹ_, it will have a tendency to turn on the back as on a
hinge, and the middle of the edge will sink deeper into the hole as it
approaches the centre of the latter. The section will therefore vary in
thickness and prove useless. This difficulty can be obviated only by the
use of a straight edge like _e f_, which will always lie in the same
plane.
[Illustration: Fig. 10.]
If the knife were perfectly rigid, and the curved edge and back lay
entirely in the same plane, this difficulty would not occur to any great
extent. But since a curved edge rarely lies wholly in the plane of the
back of the knife, and since the blades of most knives, especially those
thin ones used for cutting sections of soft tissues, yield a little to
pressure, and since this pressure is never exactly the same, it is
impossible to prevent irregularities from occurring.
While it is not necessary that the back of the knife and the cutting
edge should be parallel to each other, it is absolutely necessary that
they be in the same plane, or, as mechanics say,—“out of wind.” If this
condition be not observed, the blade will rock on the microtome table as
it passes over it, and irregularity in thickness of section will result.
A slight knowledge of geometry will enable any one to see that these
statements are strictly accurate.
As regards other points in the form of the knife, we refer to what has
been previously said when discussing the general principles which should
govern the construction of cutting tools.
We now proceed to give a few practical directions for sharpening knives
and razors and keeping them in order.
Section knives, during the process of sharpening, are subjected to three
distinct processes: 1, Grinding; 2, Honing; 3, Stropping.
Grinding is one of the most important, though it is probable that it
will rarely be undertaken by the microscopist himself. More knives and
razors are spoiled in grinding than in any other way. We have now in our
possession an excellent knife, which in an evil hour we entrusted to a
New York cutler, and received it back utterly ruined, the temper being
entirely taken out of the blades! Some of our readers may wonder at
this, but unfortunately it is too true. The cutler, to save time and
trouble, too frequently holds the blade against the stone with such
force that it becomes over-heated, especially when he is a little
careless and allows the supply of water to fall short.[20] The owner
does not find this out until he discovers that the knife is ruined,
because nothing is more easy than to cool the blade and grind off the
tell-tale blue spots. When charged with his rascality, the cutler always
denies it and lays the blame on a “soft spot in the steel”—a miserable
subterfuge—too transparent to deceive the least experienced.
Footnote 20:
Nothing is more easy than to heat a thin rod or stout wire red hot by
holding it against a dry grindstone rapidly revolving. We have often
kindled fire in this way.
The only protection against this is either to entrust the knife to a man
of known carefulness and integrity, or to stand over him while he does
it. If the knife be ground under the owner’s eye, no fear need be
entertained, because it is easy to insist upon the use of plenty of
water.
The grindstone used for section knives should be of fine grain, and it
should run true and be very straight across the face, otherwise it will
be impossible to grind the knife true, and this, as we have seen, is a
necessity.
Every microscopist that expects to do much at cutting sections must
learn to hone his own knife or razor. Of the various hones in use the
famous Turkey oilstone is said to be altogether the best. It is,
however, very difficult to get it genuine and in sufficiently large
pieces. We therefore use the German hone-slate, a softer, yellow stone
of wonderful fineness and cutting power. The ordinary whetstones are
altogether too coarse, and as for artificial stones and strops, they may
do for coarse work and common razors for _scraping_, but they will not
do for knives used for section-cutting. If any of our readers should be
so fortunate as to possess a good, old-fashioned Turkey oilstone with a
fair sized surface, let them think much of it. It is invaluable. Years
ago, when working on some investigations that required the use of very
sharp knives, we were offered, by an old English carpenter, the use of a
real Turkey stone. We have never since found anything that did its work
so quickly and so well.
To merely mention the substitutes for Turkey oilstone would fill some
pages. We therefore confine ourselves to the German hone. It is obtained
from the slate mountains in the neighborhood of Ratisbon, where it
occurs in the form of a yellow vein running vertically into the blue
slate, sometimes not more than an inch in thickness, and varying to
twelve to eighteen inches. After being quarried it is sawed into thin
slabs, which are usually cemented to a similar slab of the blue slate as
a support, and the combined stones are then set in a wooden frame and
fitted with a cover, also of wood. Unlike the Turkey oilstone, the
Arkansas stone, and some others, the German hone is soft and easily
scratched and worn. It must therefore be carefully preserved, and as the
_flatness_ of the hone is an important point, it should never be used
for anything but section knives.
Having purchased a hone, the first thing to do is to see that it is
perfectly true; that is, that its surface is a perfect plane. If hollow
or twisted, it is useless until made straight. It may be tested by means
of two steel or wood straight-edges, and the method of doing this, which
is very simple but difficult to describe, the microscopist can learn
from any cabinet maker or carpenter. If the hone be not true it must be
made so, and this is best done by grinding it with emery on a slab of
marble or, better still, a plate of cast iron which has been planed
true. (Waste castings with planed surfaces may be found in most machine
shops, and may be either borrowed or purchased for a small sum.) If the
hone is very uneven, coarse emery may be used at first; then finer must
be taken, and so on until the surface of the hone is not only true but
very smooth. This involves considerable labor, but it cannot be avoided
if a _true_ edge as well as a sharp one is desired, and hence our advice
to use the hone for nothing but section knives. Accuracy in a penknife
or a razor for shaving is of no consequence; in a section-knife it is a
_sine qua non_.
The surface of the hone must not only be true and smooth, it must also
be clean and free from dust and grit, a single particle of which may
spoil the work of hours. It should therefore, when not in use, be kept
constantly covered. It is always used with oil, and this oil should be
of a kind that does not readily dry up. Good sperm is excellent, and so
is purified neat’s-foot oil. Avoid kerosene, soap and water, and similar
fluids, which are so frequently recommended, especially under the guise
of new discoveries. They have all been tried and found wanting. After
use, the stone should be carefully wiped clean, fresh oil being applied
when it is next used. It must never be used without a liberal supply of
oil, otherwise it will become glazed and will no longer act on the
steel.
The hone being in good order, the blade is laid flat on it and moved
over its surface with circular sweeps. The method of doing this is
easily acquired, and is best learned by watching an expert cutler at
work. If we could only hit it, the perfection of this operation would be
to wear down the sides of the blade until they just met; they would then
form a keen and perfect edge. Unfortunately we cannot always hit this
point exactly; we are apt to overdo the matter just a little, and then
comes up the _bête noir_ of beginners—the “wire edge.” This is caused by
the metal of the blade bending away from the hone as soon as it becomes
of a certain degree of thinness. Once it does this it cannot be worn off
by the hone, and the only way to remove it is to draw the edge of the
knife or razor across a piece of horn or hard wood. After removing the
wire edge in this way, give the blade one or two sweeps on the hone and
then strop it. If the blade be very soft, the wire edge comes very
quickly and easily, and is very hard to get rid of. On very hard blades
the wire edge is not so apt to make its appearance. It comes, however,
on very excellent blades. It may be avoided in a measure by driving the
blade edge-first over the hone, and indeed the blade should always be
sharpened most in this way. And since the backward and forward strokes
are always nearly equal, as regards space passed over, this is
accomplished by exerting a greater pressure on the forward than on the
backward stroke.
This wire edge is a nasty thing when it breaks off on the hone. Unless
removed it will very speedily ruin a fine knife, therefore look out for
it and wipe off the hone carefully if you have any suspicion that
particles of steel have broken off the blade and got on to it.
In honing, as in everything else, however, nothing but practice will
impart skill, and he who intends to become an expert should practice on
a few old razors, grinding, honing and stropping them himself, until he
has acquired the art of giving them an edge far keener than most of the
razors ordinarily used for shaving.
The last operation to which the knife is subjected is that of stropping,
and as the blade should be stropped after every section, it becomes
important that our tools should be good and that we should know how to
use them. By having two or three knives we can always avoid grinding and
honing the blades ourselves, for it is then easy to hire the work done,
and half a dozen knives will last a careful worker a long time with
merely the aid of a good strop. But the strop he must use himself.
Fortunately no great skill is required in stropping, but the strop must
be a good one and is best home-made.
Our readers have doubtless seen the “Cheap John” strop-vendors take a
rough table knife, strop it on one of their “patent” strops and cut a
hair with it. Nevertheless such a knife, sharpened in that way, would
not cut good sections, and such a strop is not just the thing for giving
the finishing touch to a section-knife, even though the label does say
that it is “intended for surgical instruments.”[21]
Footnote 21:
The finest emery paper, glued on a strip of wood and used as a strop,
will impart to a razor an edge sufficiently keen to enable it to
remove the beard, and if paper three or four degrees coarser be used,
a very dull knife or razor may be sharpened, and afterwards stropped
on the finer emery until it is keen enough to cut a hair. But it will
be found that cutting edges produced in this way are not smooth enough
for section-cutting.
The strop should never be used with the intention of removing metal so
as to thin the edge of the knife. This is the proper function of the
hone, and those who depend most on the hone and least on the strop, will
always succeed best. The strop is used in the first place for
_smoothing_ off the edge, and in the second for removing that incipient
dullness which always comes from even the slightest use. It is true that
in the latter case the strop trespasses somewhat on the functions of the
hone, but only to a very slight extent, and stropping should never be
carried so far as to require more than a very few strokes.
[Illustration: Fig. 11.]
[Illustration: Fig. 12.]
The best material for a strop is good calf-skin, well tanned, and firmly
glued to a strip of wood with the hair side out. The leather should be
hard, otherwise it will rapidly round the edge of the knife. The way in
which this occurs is readily understood from figures 11 and 12. In Fig.
12 the leather is seen to rise up behind the edge of the blade as the
latter passes over it, so that instead of two plane facets meeting at an
angle of from 15° to 20° as left from the hone, (Fig. 12) the edge has
become considerably rounded and the ultimate angle of the cutting edge
is nearly or quite doubled. The hardest and firmest leather should
therefore be chosen, avoiding of course any leather that has been made
hard and unpliable by the action of water or other agents. Two sides
will be found enough for a strop. On one the leather should be kept
clean, while the other should be thoroughly impregnated with fine
grained, but very hard rouge or crocus.[22] Rouge is an artificial oxide
of iron prepared by exposing sulphate of iron to heat. The hardness of
the resulting powder depends upon the temperature to which it has been
exposed, and this temperature is very well indicated by the color which
the rouge assumes. Bright red or crimson rouge is soft and will not cut
steel; hard rouge, suitable for polishing steel, is purple in color, and
this quality should therefore be chosen. Great care should be taken to
see that it is free from gritty particles, and it should be well rubbed
into the leather in a dry state. All mixtures of grease and oil with
abrasive substances should be eschewed. If the leather be of good
texture and the rouge hard and fine, a very few strokes will suffice to
impart the last degree of smoothness and keenness to the blade.
Footnote 22:
Rouge may be purchased from dealers in watch-makers’ tools. Those who
cannot procure it readily may very easily prepare it for themselves.
Full directions may be found in the “Amateur’s Handbook.” New York:
Industrial Publication Company.
Since grit and dust would soon spoil the strop, it should be kept
carefully covered and protected. The rouge will require occasional
renewing, which may be done by sprinkling a little lightly over the
surface and rubbing it in with the finger. Consequently we find that the
best and most convenient holder for rouge is a bottle with some fine but
porous fabric tied over the mouth of it, forming as it were a sort of
dredge. Over the bottle mouth and its cover should be tied a paper cap
to keep out dust.
We have thus endeavored to give, in as simple a manner as possible, such
information as will guide the microscopist to the acquisition of skill
in this most important but generally despised department of his art. We
are told by the biographer of Swammerdam that a great deal of his
success was to be attributed to his skill in sharpening his fine knives
and scissors. The same is also stated of other noted workers, and yet
none give even the slightest directions to the novice who desires to
follow in their footsteps. There is not to our knowledge a single book
in the English language which treats fully of this subject, with perhaps
the exception of Holtzapffel’s work on “Mechanical Manipulation,” and
this has long been out of print. As previously stated, however, it is
only by practice that the necessary finger-skill can be acquired, but
this skill is well worth acquiring at any cost. The man who depends upon
cutlers and knife grinders will never make satisfactory progress.
NOTE H.
We are inclined to believe that the difficulty of enclosing glycerine
does not arise from the dissolving action of this liquid, so much as
from the great adhesiveness which exists between it and glass. Oily
cements are of course attacked by glycerine, but shellac and several
other gums are insoluble, and drying oils, if well oxidized are also
insoluble. Where cells are used they must be first firmly attached to
the clean slide, and the cement should also be brought into direct
contact with the clean cover. Glycerine does not evaporate at ordinary
temperatures, and consequently wherever the slide or cover is soiled
with it, no cement will ever adhere until the glycerine has been wiped
off. There is no prospect of its drying.
NOTE I. Page 48.
The solution here referred to, and which is known as Kleinenberg’s
solution, is made in the following way:
(1.) Make a saturated solution of crystallized calcium chloride in 70
per cent. alcohol, and add alum to saturation. (2.) Make also a
saturated solution of alum in 70 per cent. alcohol. Add (1) to (2) in
the proportion of 1:8. To the mixture add a few drops of a saturated
solution of hæmatoxylin in absolute alcohol.
This solution may be used in very many cases for staining sections, in
place of the ordinary watery solution of logwood alum. It may, if
required, be diluted with the mixture of 1 and 2. The stained sections
are placed at once in strong spirit.
------------------------------------------------------------------------
INDEX.
Acetic acid, 38
Adipose tissue, 63
Æther microtome, 24
Air-bubbles, 40, 55, 62
Alcohol, 17, 19
absolute, 54
Balsam, Canada, 46, 51
to clean from slide, 55
Beale’s carmine fluid, 33
Bell’s cement, 41
Blue staining fluid, 60
Bone, 16, 57
decalcified, 58
Bottles for media, &c., 48
Brain, 59
Brunswick black, 41
Capillary attraction, 55
Carmine staining, 32
Carpenter, Dr., 37, 41
Cartilage, 60
Cells, to make, 59
Centring slide, 38
Chloroform-balsam, 51, 54
Chromic acid, 17
Clove oil, 51, 54
Coffee berry, 62
Cover, applying, 37
German plan, 39
Crochet-needle holders, 39
Double staining, 73
Dry mounting, 58, 65, 72
Fat, 63
Finishing slide, 54
Fibro-cartilage, 62
Freezing method, 42-47
Fresh tissues, to cut, 15-17
Fruit stones, 58
Gardner, Mr., 25, 31
Gelatine cement, 41
Glycerine, 37, 38
Gold staining, 58
Gum, imbedding in, 22, 66
strong, 43
weak, 31, 71
Hair, 63-69
Hæmatoxylon, _see_ Logwood
Hand-sections, 19
Hardening agents, 19
Hoggan, F. Elizabeth, Dr., 61
Hone, 26, 57
Horn, hoofs, &c., 16, 64
Hyaline cartilage, 62
Ice, for freezing, 43
Injections, to harden, 19
Ink staining, 70
Intestine, 42, 43, 64
Iron staining, 61
Kay, J. A., Mr., 52
Kidney, 17
Klein’s _lifter_, 35
Knife, section, 25, 30
Leaves, &c., to cut, 12
Ligaments, intervertebral, 62
Lime, carbonate of, 37
Liver, 65
Logwood staining, 47
Lung, 43, 65
Martindale’s logwood fluid, 47
Media, mounting, 38
Methylated spirit, _see_ Alcohol
Microtome, 21
choice of, 22
Microtome, employment of, 29
freezing, 43, 44
imperfection in, 23, 46
Rutherford’s, 31
Muscle, 66
Needles, microscopical, 39
Orange-peel, 67
Ovary, 67
Papillæ of finger, 69
Paraffine, adhering to sections, 27
imbedding in, 20, 26
mixture, 26
_rising_ in tube, 23
shrinks in cooling, 29
Paul, Dr., 70
Pathological specimens, 19, 62
Picro-carmine, 59, 68, 72
Porcupine quill, 67
Potato, 67
Preparation of animal tissues, 16
vegetable tissues, 15
Prussian blue pigment, 38
Rangoon oil, 14, 47
Razors, 20, 26, 29, 30
Rush, 78
Rutherford’s microtome, 24, 31
“Practical Histology,” 43
Salicylic acid, 44
Salt, for freezing, 45
Schäfer’s “lifter,” 35
“Practical Histology,” 48
Scissors, section, 13
Sebaceous glands, 69
Section-cutter, _see_ Microtome
Section-knife, 24
Sections, thickness of, 31
to dehydrate, 52
to transfer, 33
Seeds, 63
Skin, 68
Sperm oil, 44
Spinal cord, 69
Spirit lamp for balsam, 53
weak, 36, 59
Sponge, 71
Spoon, section, 35, 54
Staining agents, 31
Strop, razor, 26, 29
Stomach, 71
Sudoriferous glands, 69
Taste-bulbs, 71
Teeth, 16, 58
Tongue, 71
Tubes, collapsible, 56
Valentin’s knife, 14
Vegetable ivory, 72
tissues, to prepare, 12
Vessels, porcelain, 33
Water-bath, cheap, 29
Whalebone, 64
White zinc cement, 56
Wood, 72
------------------------------------------------------------------------
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HOW TO USE THE MICROSCOPE.—How to Care for the Microscope.
HOW TO COLLECT OBJECTS.—Where to find Objects.—What to Look for.—How to
Capture Them.—Nets.—Bottle-Holders.—Spoons.—New Form of Collecting
Bottle.—Aquaria for Microscopic Objects.—Dipping Tubes.
THE PREPARATION AND EXAMINATION OF OBJECTS.—Cutting Thin Sections
of Soft Substances.—Sections of Wood and Bone.—Improved
Section-Cutter.—Sections of Rock.—Knives.—Scissors.—Needles.—Dissecting
Pans and Dishes.—Dissecting Microscopes.—Separation of Deposits from
Liquids.—Preparing whole Insects.—Feet, Eyes, Tongues, Wings, etc.,
of Insects.—Use of Chemical Tests.—Liquids for Moistening
Objects.—Refractive Power of Liquids.—Covers for Keeping out
Dust.—Errors in Microscopical Observations.
PRESERVATION OF OBJECTS.—General Principles.—Recipes for Preservative
Fluids.—General Rules for Applying them.
MOUNTING OBJECTS.—Apparatus and Materials for: Slides, Covers, Cells,
Turn-Table, Cards for Making Cells, Hot-Plate, Lamps, Retort Stand,
Slide-Holder, Mounting Needles, Cover Forceps, Simple Form of Spring
Clip, Centering Cards, Gold-Size, Black Japan, Brunswick Black, Shellac,
Bell’s Cement, Sealing Wax Varnish, Colored Shellac, Damar Cement,
Marine Glue, Liquid Glue, Dextrine.—Mounting Transparent Objects
Dry.—Mounting in Balsam.—Mounting in Liquids.—Mounting of Whole
Insects.—How to Get Rid of Air-Bubbles.—Mounting Opaque Objects.
FINISHING THE SLIDES.
May be obtained from any Bookseller or News Agent, or will be sent by
mail, postage paid, on receipt of price.
THE INDUSTRIAL PUBLICATION COMPANY,
P. 0. Box 4875. 176 Broadway, New York.
------------------------------------------------------------------------
_GENESIS AND GEOLOGY._
--------------
THE CHEMICAL HISTORY
OF
The Six Days of Creation.
BY JOHN PHIN, C. E.,
EDITOR OF “THE AMERICAN JOURNAL OF MICROSCOPY.”
_1 Vol., 12mo._ _Cloth._ _75 cents._
--------------
The following are a few of the Opinions of the Press:
This is a small book, but full of matter. * * * We think this book is
full of interest and value; and as the discussions concerning the
harmony of science and faith are rife at the present day, we commend the
reasonings of Mr. Phin to the great number of readers and students who
are investigating these subjects.—_The Presbyterian_ (Philadelphia).
The author gives a new solution of this difficult question, and
certainly presents many very plausible arguments in support of his
theory.—_Sunday-School Workman._
A very candid and ingenious essay.—_Christian Union_ (H. W. Beecher’s
paper).
No one can read this book without compensation, without becoming more
thoughtful concerning the phenomena of creation; and he need lose none
of his reverence for the supremacy of the Divine Law.—_Rural New
Yorker._
We could say much in commendation of Prof. Phin’s little book. An
intelligent reader can hardly fail to be interested in it, and many
might be benefited.—_Country Gentleman._
The book can not fail to interest even those who do not fully accept the
theory it advocates.—_Boston Journal of Chemistry._
It is a new scientific view of the matter.—_Phrenological Journal._
The book, although not large, will prove exceedingly interesting to all
who have ever directed attention to this matter, and contains more solid
and suggestive thought than many voluminous treatises on the
subject.—_Insurance Monitor._
The work is ingenious and original, and presents many striking
suggestions.—_American Baptist._
We believe Prof. Phin has started upon the correct basis, and his theory
is mainly tenable. His views are presented in a manner which, though
terse, is easily comprehended.—_Daily Guardian_ (Paterson, N. J.)
------------------------------------------------------------------------
THE YOUNG SCIENTIST,
A Practical Journal for Amateurs.
=ISSUED MONTHLY. Price 50 Cents per year.=
It is characteristic of young Americans that they want to be DOING
something. They are not content with merely _knowing_ how things are
done, or even with _seeing_ them done; they want to do them themselves.
In other words, they want to experiment. Hence the wonderful demand that
has sprung up for small tool chests, turning lathes, scroll saws, wood
carving tools, telegraphs, model steam engines, microscopes and all
kinds of apparatus. In nine cases out of ten, however, the young workman
finds it difficult to learn how to use his tools or apparatus after he
has got them. It is true that we have a large number of very excellent
text-books, but these are not just the thing. What is wanted is a living
teacher. Where a living teacher cannot be found, the next best thing is
a live journal, and this we propose to furnish. And in attempting this,
it is not our intention to confine ourselves to mere practical
directions. In these days of knowledge and scientific culture, the “Why”
becomes as necessary as the “How.” The object of the YOUNG SCIENTIST is
to give clear and easily followed directions for performing chemical,
mechanical and other operations, as well as simple and accurate
explanations of the principles involved in the various mechanical and
chemical processes which we shall undertake to describe.
The scope and character of the journal will be better understood from an
inspection of a few numbers, or from the list of contents found on a
subsequent page, than from any labored description. There are, however,
three features to which we would call special attention:
CORRESPONDENCE.—In this department we intend to place our readers in
communication with each other, and in this way we hope to secure for
every one just such aid as may be required for any special work on hand.
EXCHANGES.—An exchange column, like that which has been such a marked
success in the _Journal of Microscopy_, will be opened in the Young
Scientist. Yearly subscribers who may wish to _exchange_ tools,
apparatus, books, or the products of their skill, can state what they
have to offer and what they want, _without charge_. Buying and selling
must, of course, be carried on in the advertising columns.
ILLUSTRATIONS.—The journal will make no claims to the character of a
“picture book,” but wherever engravings are needed to make the
descriptions clear they will be furnished. Some of the engravings which
have already appeared in our pages are as fine as anything to be found
in the most expensive journals.
Special Notice.
As our journal is too small and too low-priced to claim the attention of
news dealers, we are compelled to rely almost wholly upon subscriptions
sent directly to this office. As many persons would no doubt like to
examine a few numbers before becoming regular subscribers, we will send
four current numbers as a trial trip for
FIFTEEN CENTS.
--------------
CLUBS.
Where three or more subscribe together for the journal, we offer the
following liberal terms:
3 copies for $1.25
5 〃 〃 2.00
7 〃 〃 2.75
10 〃 〃 3.50
Advertisements, 30 cents per line.
As postal currency has nearly disappeared from circulation, we receive
postage stamps of the lower denominations (ones, twos and threes) at
their full value. Postal orders are, however, much safer and more
convenient. To avoid delay and mistakes address all communications to
“THE YOUNG SCIENTIST, Box 4875, New York,” and make all checks and
orders payable to John Phin.
------------------------------------------------------------------------
=WHAT PEOPLE SAY OF US.=
In a letter to the Editor, Oliver Wendell Holmes, the genial “Autocrat
of the Breakfast Table,” says: “I am much pleased with the YOUNG
SCIENTIST. It makes me want to be a boy again.”
“It is a little publication, calculated to call out and educate all the
latent ingenuity and thirst for knowledge which the youthful mind
possesses, and we hope it will win its way into every household in the
land.”—[Scientific Press.
“We have never seen a periodical, designed for youth, which came nearer
to our ideal of what such a journal should be.”—[Canadian Pharmaceutical
Journal.
“The YOUNG SCIENTIST is one of the choicest publications for juvenile
minds in this country. Every page treats on subjects of importance to
young and old, portrayed in a clearly comprehensive manner, which at
once interests the young idea in its careful perusal.”—[Lapeer Clarion.
“It seems to fill the bill.”—[Newport Daily News.
“It is pleasing to note that its youthful subscribers will not be misled
by clap-trap advertisements or advertisements of patent medicines, which
will not be received at any price. The YOUNG SCIENTIST is doing good
work in setting its face against this class of humbugs.”—[Manufacturing
and Trade Review.
“The work is a copiously illustrated monthly, and is full of practical
hints that will instruct and amuse the young folks.”—[Industrial School
Advocate.
“A small but elegant and very instructive monthly.”—[Pittsburg
Chronicle.
“Contains the best possible reading for the young of both
sexes.”—[Ottawa Journal.
“We can safely recommend this magazine as one of the very best
publications for the young folks.”—[The Independent, Fenton, Mich.
“This journal occupies a new field, and is needed to put the minds of
our youth on the right track to secure a correct understanding of the
nature of things.”—[Wayland Press.
“It is ably edited by John Phin, who will make a large place in the
heart of the rising generation, if he persists in his venture. We hope
his success in the field will be equal to the article furnished—first
best.”—[Sunset Chimes.
“The articles are written in a popular, readable style, and profusely
illustrated.”—Akron City Times.
“The YOUNG SCIENTIST is excellent in conception, and well designed to
amuse and instruct young people.”—[Chicago Evening Journal.
“The YOUNG SCIENTIST is a handsome monthly magazine, each number
containing about 16 pages, handsomely illustrated. It will supply a
place which has been heretofore unoccupied. The copy before us comes
fully up to the promise of the prospectus.”—[The Times, Iroquois, Mich.
“It is a journal which should be in the hands of both young and old, and
is a great benefit to the young scientist as well as the advanced
professor. It is a thousand times more valuable than the dime novel
series, so much read by boys. Parents would do well to have it in their
households.”—[The Iron Home.
“This publication is a new launch, and it is very gratifying to witness
the ableness which pervades its pages.”—Amherst Free Press.
------------------------------------------------------------------------
THE AMERICAN
JOURNAL OF MICROSCOPY,
AND
_POPULAR SCIENCE_.
--------------
PROSPECTUS.
The object of the JOURNAL OF MICROSCOPY is to diffuse a knowledge of the
best methods of using the Microscope; of all valuable improvements in
the instrument and its accessories; of all new methods of microscopical
investigation, and of the most recent results of microscopical research.
The JOURNAL does not address itself to those who have long pursued
certain special lines of research, and whose wants can be supplied only
by elaborate papers, which, from their thoroughness, are entitled to be
called monographs rather than mere articles. It is intended rather to
meet the wants of those who use the microscope for purposes of general
study, medical work, class instruction, and even amusement, and who
desire, in addition to the information afforded by text-books, such a
knowledge of what others are doing as can be derived only from a
periodical. With this object in view, therefore, the publishers propose
to make the JOURNAL so simple, practical and trustworthy, that it will
prove to the advantage of every one who uses the microscope at all to
take it.
ILLUSTRATIONS.—The JOURNAL will be freely illustrated by engravings
representing either objects of natural history or apparatus connected
with the microscope.
TRANSACTIONS OF SOCIETIES.—THE AMERICAN JOURNAL OF MICROSCOPY is not the
organ of any Society, but it gives the proceedings of all Societies
whose officers send us a report. As the JOURNAL is devoted _wholly_ to
Microscopy, and is in good form and size for binding, no better medium
can be had for preserving the scientific records of any society. Matters
of mere business routine we are frequently obliged to omit for want of
room.
EXCHANGES.—An important feature of the JOURNAL is the exchange column,
by means of which workers in different parts of the country are enabled,
without expense, except for postage, to exchange slides and materials
with each other.
=TERMS.=
During the first two years of its existence, the subscription to the
AMERICAN JOURNAL OF MICROSCOPY was only fifty cents per year, but at the
request of more than two-thirds of the subscribers, the size of the
JOURNAL has been doubled, and the price raised to
=ONE DOLLAR PER YEAR.=
Four copies for three dollars. Those who wish to economize in the
direction of periodicals, would do well to examine our clubbing list.
FOREIGN SUBSCRIBERS.—The JOURNAL will be sent, postage paid, to any
country in the Postal Union for $1.24, or 5 shillings sterling per year.
English postage stamps, American currency or American postage stamps
taken in payment. In return for a postal order or draft for £1 5s., five
copies of the JOURNAL will be furnished and mailed to different
addresses. Make all drafts and postal orders payable to John Phin.
BACK VOLUMES.—We have on hand a few copies of Vols. I and II, bound in
handsome cloth cases, which we offer for $1 25 each. Vols. I and II,
bound, and the numbers of Vol. III, as issued, we offer for $2.50. We
can no longer supply complete sets of 1876-7 in sheets. To those who
wish to examine the journal, we will send ten odd numbers for 25 cents.
=Advertisements.=
The JOURNAL OF MICROSCOPY, from its very nature, is a visitor to the
very best families, and its value as an advertising medium has therefore
proved to be much above that of average periodicals. A few select
advertisements will be inserted at the rate of 30 cents per line,
nonpariel measure, of which twelve lines make an inch. Address
AMERICAN JOURNAL OF MICROSCOPY,
P. O. Box 4875, New York.
------------------------------------------------------------------------
● Transcriber’s Notes:
○ Missing or obscured punctuation was corrected.
○ Typographical errors were silently corrected.
○ Inconsistent spelling was made consistent when a predominant form
was found in this book; otherwise it was not changed.
○ Forms of the name Schäfer were regularized to match the title page
of his “A Course of Practical Histology.”
○ Text that was in italics is enclosed by underscores (_italics_);
text that was bold by “equal” signs (=bold=).
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