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BIOLOGY
UBRAPV
THE MICROSCOPIST.
ZENTMAYER'S LARGEST MICROSCOPE.
• ONE-THIRD ACTUAL SIZE.
THE
MICROSCOPIST
MANUAL OF MICROSCOPY
AND
COMPENDIUM OF THE MICROSCOPIC SCIENCES,
MICRO-MINERALOGY, MICRO-CHEMISTRY, BIOLOGY, HISTOLOGY,
AND PATHOLOGICAL HISTOLOGY.
THIRD EDITION.
KEWKITTEN AND GEEATLY ENLAKGED.
WITH
TWO HUNDRED AND FIVE ILLUSTRATIONS.
J. H. WYTHE, A.M., M.D.,
PROFESSOR OF MICROSCOPY AND BIOLOGY IN THE MEDICAL COLLEGE OF THE PACIFIC,
SAN FRANCISCO.
PHILADELPHIA:
LINDSAY & BLAKISTON.
1877.
BIOLOGY
UVRARY
Entered according to Act of Congress, in the year 1877,
By LINDSAY & BLAKISTON,
In the office of the Librarian of Congress at Washington, D. C.
/ 3.
PHILADELPHIA:
SHERMAN & CO., PRINTERS.
RESPECTFULLY DEDICATED
TO THE -
SAN FRANCISCO MICROSCOPICAL SOCIETY,
AS A TESTIMONY
TO THE
ZEAL AND INDUSTRY OF ITS MEMBERS
IN THE PROSECUTION
OF
MICROSCOPIC SCIENCE.
PREFACE.
THE progress of microscopic science may be well illus-
trated by a comparison between the present and former
editions of this book. The author's intention was to
place within the reach of the student of nature a com-
pendium of microscopy, free from unnecessary verbiage,
which should aid in every department of natural science.
It is 110 small compliment to such a work that for a
quarter of a century it should hold a place among works
of reference, although surrounded by larger and more
pretentious volumes. In order to meet the request of
the publishers for another edition, it has been found
necessary to rewrite the entire book, and although the
original design has been kept in view, the numerous
additions to our science render considerable enlargement
needful, notwithstanding the effort made to concentrate
the material into the smallest compass consistent with
perspicuity.
The vision of microscopy sweeps over all the world,
and embraces all forms of organic and inorganic ex-
Vlll PREFACE.
istence. To give directions respecting most approved
methods, and to classify the most important facts, has
required labor, which it is hoped will result in rendering
the work a necessary companion to the student, and an
aid to the progress of real science.
Many of the figures illustrating the lower forms of life,
and normal and pathological histology, have been drawn
from the works of Carpenter, Beale, Frey, Strieker, Bill-
roth, and Rindfleisch, to which the more advanced stu-
dent is referred for further details.
January, 1877.
CONTENTS.
CHAPTER I.
HISTORY AND IMPORTANCE or MICROSCOPY.
Application of the Microscope to Science and Art — Progress of Micros-
copy, . . 17-21
CHAPTER II.
THE MICROSCOPE.
The Simple Microscope — Chromatic and Spherical Aberration— Com-
pound Microscope — Achromatic Object-glasses — Eye-pieces — Me-
chanical Arrangements — Binocular Microscope, . . . 21-32
CHAPTER III.
MICROSCOPIC ACCESSORIES.
Diaphragms — Condensers — Oblique Illuminators — Dark-ground ditto —
Illumination of Opaque Objects — Measuring and Drawing Objects —
Standards of Measurement — Moist Chamber — Gas Chamber — Warm
Stage — Polari scope — Microspectroscope — Nose-piece — Object-finders
— Micro-photography, ........ 32-48
CHAPTER IV.
USE OP THE MICROSCOPE.
Care of the Instrument — Care of the Eyes— Table— Light— Adjustments
— Errors of Interpretation — Testing the Microscope, . . 48-58
B
X CONTENTS.
CHAPTER V.
MODERN METHODS OF EXAMINATION.
Preliminary Preparation of Objects— Minute Dissection— Preparation
of Loose Textures— Preparation by Teasing— Preparation by Section
—Staining Tissues— Injecting Tissues— Preparation in Vi>cid Media
—Fluid Media— Indifferent Fluids— Chemical Reagents— Staining
Fluids Injecting Fluids — Preservative Fluids— Cements, . 58-76
CHAPTER VI.
MOUNTING AND PRESERVING MICROSCOPIC OBJECTS.
Opaque Objects— Cells — Dry Objects — Mounting in Balsam or Dammar —
Mounting in Fluid — Cabinets — Collecting Objects — Aquaria, 76-83
CHAPTER VII.
THE MICROSCOPE IN MINERALOGY AND GEOLOGY.
Preparation of Specimens — Examination of Specimens — Crystalline
Forms — Crystals witbin Crystals— Cavities in Crystals — Use of Po-
larized Light — Origin of Rock Specimens — Materials of Organic
Origin — Microscopic Paleontology, ..... 84-98
CHAPTER VIII.
THE MICROSCOPE IN CHEMISTRY.
Apparatus and Modes of Investigation — Preparation of Crystals for the
Polariscope — Use of the Microspectroscope — Inverted Microscope —
General Micro-chemical Tests — Determination of Substances — Al-
kalies— Acids — Metallic Oxides — Alkaloids — Crystalline Forms of
Salts, 98-115
CHAPTER IX.
THE MICROSCOPE IN BIOLOGY.
Theories of Life — Elementary Unitor Cell — Cell-structureand Formation
— Phenomena of Bioplasm — Movements of Cells — Microscopic Dem-
onstration of Bioplasm — Chemistry of Cells and their Products — Va-
rieties of Bioplasm — Cell-genesis — Reproduction in Higher Organ-
isms— Alternation of Generations — Parthenogenesis —Transforma-
tion and Metamorphosis — Discrimination of Living Forms, 116-127
CONTENTS. xi
CHAPTER X.
THE MICROSCOPE IN VEGETABLE HISTOLOGY AND BOTANY.
Molecular Coalescence — Cell-substance in Vegetables — Cell- wall or Mem-
brane— Ligneous Tissue— Spiral Vessels — Laticiferous Vessels — Si-
liceous Structures — Formed Material in Cells — Forms of Vegetable
Cells — Botanical Arrangement of Plants — Fungi — Protopbytes —
Desmids — Diatoms — Nostoc — Oscillatoria — Examination of the
Higher Cryptogamia — Examination of Higher Plants, . 128-157
CHAPTER XI.
THE MICROSCOPE IN ZOOLOGY.
Monera — Rhizopods — Infusoria — Rotatoria — Polyps — Hydroids — Aca-
lephs — Echinoderms — Bryozoa — Tunicata — Conchifera — Gastero-
poda — Cephalopoda — Entozoa — Annulata — Crustacea — Insects —
Arachnida — Classification of the Invertebrata, . . . 158-182
CHAPTER XII.
THE MICROSCOPE IN ANIMAL HISTOLOGY.
Histo-chemistry — Histological Structure — Simple Tissues — Blood —
Lymph and Chyle — Mucus — Epithelium — Hair and Nails — Enamel
— Connective Tissues — Compound Tissues — Muscle — Nerve — Glan-
dular and Vascular Tissue— Development of the Tissues — Digestive
and Circulatory Organs — Secretive Organs — Respiratory Organs —
Generative Organs — Locomotive Organs — Sensory Organs — Organs
of Special Sense — Suggestions for Practice, . . . 182-226
CHAPTER XIII.
THE MICROSCOPE IN PRACTICAL MEDICINE AND PATHOLOGY.
Microscopic Appearances after Death of the Tissues — Morbid Action in
Tissues — New Formations — Examination of Urinary Deposits —
Human Parasites— Examination of Sputa— Microscopic Hints in
Materia Medica and Pharmacy, ..... 226-245
1 1 B R A R Y
NIVERSITY OF
THE MICROSCOPIST.
CHAPTER I.
HISTORY AND IMPORTANCE OF MICROSCOPY.
THE term microscopy, meaning the use of the micro-
scope, is also applied to the knowledge obtained by this
instrument, and in this sense is commensurate with a
knowledge of the minute structure of the universe, so far
as it may come under human observation. Physics and
astronomy treat of the general arrangement and motions
of masses of matter, chemistry investigates their constitu-
tion, and microscopy determines their minute structure.
The science of histology, so important to anatomy and
physiology, is wholly the product of microscopy, while
this latter subject lends its aid to almost every other
branch of natural science.
To the student of physical phenomena this subject un-
folds an amazing variety developed from most simple
beginnings, while to the Christian philosopher it gives the
clearest evidence of that Creative Power and Wisdom
before whom great and small are terms without meaning.
In the arts, as well as in scientific investigations, the
microscope is used for the examination and preparation of
delicate wrork. The jeweller, the engraver, and the miner
find a simple microscope almost essential to their employ-
ments. This application of the magnifying power of lenses
was known to the ancients, as is shown by the glass lens
2
18 THE MICROSCOPIST.
found at Nineveh, and by the numerous gems and tablets
so finely engraved as to need a magnifying glass to detect
their details.
In commerce, the microscope has been used to detect
adulterations in articles of food, drugs, and manufactures.
In a single year $60,000 worth of adulterated drugs was
condemned by the New York inspector, and, so long as
selfishness is an attribute of degraded humanity, so long
will the microscope be needed in this department.
In agriculture and horticulture microscopy affords valu-
able assistance. It has shown us that mildew and rust in
wheat and other food-grains, the " potato disease," and
the "vine disease," are dependent on the growth of minute
parasitic fungi. It has also revealed many of the minute
insects which prey upon our grain-bearing plants and fruit
trees. The damage wrought by these insects in the United
States alone has been estimated by competent observers
as not less than three hundred millions of dollars in each
year. The muscardine, which destroys such large num-
bers of silk-worms in France and other places, is caused
by a microscopic fungus, the Botrytis bassiana.
The mineralogist determines the character of minute
specimens or of thin sections of rock, and the geologist
finds the nature of many fossil remains by their magnified
image in the microscope.
The chemist recognizes with this instrument excessively
minute quantities and reactions which would otherwise
escape observation. Dr. Wormley shows that micro-
chemical analysis detects the reaction of the 10,000th to
the 100,000th part of a grain of hydrocyanic acid, mer-
cury, or arsenic, and very minute quantities of the vege-
table alkaloids may be known by a magnified view of their
sublimates. The micro-spectroscope promises still more
wonderful powers of analysis by the investigation of the
absorption bands in the spectra of different substances.
In biology the wonderful powers of the microscope find
HISTORY AND IMPORTANCE OF MICROSCOPY. 19
their widest range. If we see not life itself, we see its
first beginnings, and the process of its development or
manifestation. If we see not Nature in her undress, we
trace the elementary warp and woof of her mystic drapery.
In vegetable and animal physiology we see, by its
means, not only the elementary unit — the foundation-stone
of the building — but also chambers and laboratories in
the animated temple, which we should never have sus-
pected— tissues and structures not otherwise discoverable
— not to speak of species innumerable which are invisible
to the naked eye.
In medical science and jurisprudence the contributions
of microscopy have been so numerous that constant study
in this department is needed by the physician who would
excel or even keep pace with the progress of his profes-
sion. Microscopy may be truly called the guiding genius
of medical science.
Even theology has its contribution from microscopy.
The teleological view of nature, which traces design, re-
ceives from it a multitude of illustrations. In this de-
partment the war between skeptical philosophy and theol-
ogy has waged most fiercely; and if the difference between
living and non-living matter may be demonstrated by the
microscope, as argued by Dr. Beale and others, theology
sends forth a paean of victory from the battlements of this
science.
The attempts made by early microscopists to determine
ultimate structure were of but little value from the im-
perfections of the instruments employed, the natural mis-
takes made in judging the novel appearances presented,
and the treatment to which preparations were subjected.
In late years the optical and mechanical improvements in
microscopes have removed one source of error, but other
sources still remain, rendering careful attention to details
and accurate judgment of phenomena quite essential. Care-
ful manipulation and minute dissection require a knowledge
20 * THE MICROSCOPIST.
of the effects of various physical and chemical agencies, a
steady hand, and a quick-discerning eye. Above all,
microscopy requires a cultured mind, capable of readily
detecting sources of fallacy, and such a love of truth as
enables a man to free himself from all preconceived no-
tions of structure and from all bias in favor of particular
theories and analogies. What result is it possible to draw
from the observations of those who boil, roast, macerate,
putrefy, triturate, and otherwise injure delicate tissues,
except for the purpose of isolating special structures or
learning the effects of such agencies ? Yet many of the
phenomena resulting from such measures have been de-
scribed as primary, and theories of development have been
proposed on the basis of such imperfect knowledge.
Borelli (1608-1656) is considered to be the tirst who
applied the microscope to the examination of animal
structure. Malpighi (1661) first witnessed the actual cir-
.culation of the blood, which demonstrated the truth of
Harvey's reasoning. He also made manj^ accurate obser-
vations in minute anatomy. Lewenhoeck, Swammerdam,
Lyonet, Lieberkuhn, Hewson, and others, labored also in
this department. When we remember that these early
laborers u^ed only simple microscopes, generally of their
own construction, we must admire their patient industry,
skilful manipulation, and accurate judgment. In these
respects they are models to all microscopists.
Within the last quarter of a century microscopic ob-
servers may be numbered by thousands, and some have
attained an eminent reputation. At the present day, in
Germany, England, France, and the United States, the
most careful and elaborate investigations are being made,
older observations are repeated and corrected, new discov-
eries are rapidly announced, and the most hidden recesses
of nature are being explored.
It is proposed in this treatise to give such a resume of
microscopy as shall enable the student in any department
THE MICROSCOPE. 21
to pursue original investigations with a general knowl-
edge of what has been accomplished by others. To this
end a comprehensive view of the necessary instruments
and details of the art, or what the Germans call technol-
ogy, is first given, and then a brief account of the appli-
cation of the microscope to various branches of science,
especially considering the needs of physicians and stu-
dents of medicine.
CHAPTER II.
THE MICROSCOPE.
The Simple Microscope. — The magnifying power of a
glass lens (from lens, a lentil ; because made in the shape
of its seeds) was doubtless known to the ancients, but only
in modern times has it been applied in scientific research.
The forms of lenses generally used are the double convex,
with two convex faces ; piano convex, with one face flat
and the other convex ; double concave, with two concave
faces ; plajio-concave, with one flat and one concave face ;
and the meniscus, with a concave and a convex face.
In the early part of the seventeenth century very mi-
nute lenses were used, and even small spherules of glass.
Many of the great discoveries of that period were made
by these means. A narrow strip of glass was softened in
the flame of a spirit-lamp and drawn to a thread, on the
end of which a globule wras melted and placed in a thin
folded plate of brass, perforated so as to admit the light.
Some of these globules were so small as to magnify sev-
eral hundred diameters. Of course, they were inconve-
nient to use, and larger lenses, ground on a proper tool,
were more common.
The magnifying power of lenses depends on a few simple
22 THE MICROSCOPIST.
optical laws, concerning refraction of light, allowing the
eye to see an object under a larger visual angle ; so that
the power of a simple microscope is in proportion to the
shortness of its focal length, or the distance from the lens
to the point where a distinct image of the object is seen.
This distance may be measured by directly magnifying
an object with the lens, if it be a small one, or by casting
an image of a distant window, candle, etc., upon a paper
or wall. The focus of the lens is the point where the
image is most distinct. Different persons see objects
naturally at different distances, but ten inches is consid-
ered the average distance for the minimum of distinct
vision. A lens, therefore, of two inches focal length,
magnifies five diameters ; of one inch focus, ten diameters ;
of one-half inch, twenty diameters ; of one-eighth inch,
eighty diameters ; etc.
Simple microscopes are now seldom used, except as
hand magnifiers, or for the minute dissection and prepa-
ration of objects. They are used for the latter purpose,
when suitably mounted with a convenient arm, mirror,
etc., because of the inconvenience of larger and otherwise
more perfect instruments.
Single lenses, of large size, are also used for concentra-
ting the light of a lamp on an object during dissection, or
on an opaque object on the stage of a compound micro-
scope.
There are imperfections of vision attending the use of
all common lenses, arising from the spherical shape of the
surface of the lens, or from the separation of the colored
rays of light when passing through such a medium.
These imperfections are called respectively spherical and
chromatic aberration. To lessen or destroy these aberra-
tions, various plans have been proposed by opticians. For
reducing spherical aberration, Sir John Herschel pro-
posed a doublet of two plano-convex lenses, whose focal
lengths are as 2.3 to 1, with their convex sides together;
THE MICROSCOPE. 23
and Mr. Coddington invented a lens in the form of a
sphere, cut awray round the centre so as to assume the
shape of an hour-glass. This latter, in a convenient set-
ting, is one of the best pocket microscopes. Dr. Wollas-
ton's doublet consists of two plano-convex lenses, whose
focal lengths are as 1 to 3, with the plane sides of each
and the smallest lens next the object. They should be
FIG. 1.
Holland's Triplet.
about the difference of their focal lengths apart, and a
diaphragm or stop — an opaque screen with a hole in it —
placed just behind the anterior lens. This performs ad-
mirably, yet has been further improved by Mr. Holland
by making a triplet of plano-convex lenses (Fig. 1), with
the stop between the upper lenses.
The Compound Microscope consists essentially of two
convex lenses, placed some distance apart, so that the
image made by one may be magnified by the other.
These are called the object-glass and the eye-glass. In
Fig. 2, A is the object-glass, which forms a magnified
image at c, which is further enlarged by the eye-glass B.
An additional lens, D, is usually added, to enlarge the
field of view. This is called the field-glass. Its office, as
in the figure, is to collect more of the rays from the
object-glass and form an image at F, which is viewed by
the eye-glass.
Owing to chromatic aberration, an instrument of this
kind is still imperfect, presenting rings of color round the
edge of the field of view as well as at the edge of the
magnified image of an object, together with dimness and
2-4
THE MICROSCOPIST.
confusion of vision. This may be partly remedied by a
small hole or stop behind the object-glass, which reduces
the aperture to the central rays alone, yet it is still un-
FIG. 2.
Compound Microscope.
satisfactory. Some considerable improvement may result
from using Wollaston's doublet as an object-glass, but the
THE MICROSCOPE.
25
achromatic object-glasses now supplied by good opticians
leave nothing to be desired.
Object-glasses. — A general view of an achromatic object-
glass is given in Fig. 3. It is a system of three pairs of
lenses, 1, 2, 3, each composed of a double convex of crown
glass and a plano-concave of flint, a, 6, c, represents the
angle of aperture, or the cone of rays admitted. It is
unnecessary to consider the optical principles which un-
derlie this construction. Different opticians have different
formulae and propose various arrangements of lenses, and
there is room for choice among the multitude of micro-
scopes presented for sale. For high powers, the German
FIG. 3.
FIG. 4.
pss^^^^^l
Achromatic Object-glass.
Huygenian Eye-piece.
and French opticians have lately proposed a principle of
construction which is known as the immersion system.
It consists in the interposition of a drop of water between
the front lens of the objective and the covering glass over
the object. This form of object-glass is corning into gen-
eral use. For the more perfect performance of an objec-
tive, it is necessary that it should be arranged for correct-
ing the effect of different thicknesses of covering glass.
This is accomplished by a fine screw movement, which
brings the front pair of lenses (1, Fig. 3) nearer or further
from the object. In this way the most distinct and accu-
rate view of an object may be obtained.
26 THE MICROSCOPIST.
Eye pieces. — The eye-piece usually employed is the Huy-
genian, or negative eye-piece (Fig. 4). This is composed
of two plano-convex lenses, with their plane sides next
the eye. Their focal lengths are as 1 to 3, and their
distance apart half the sum of their focal distances.
Several of these, having different magnifying powers, are
supplied with good microscopes. It is best to use a weak
eye-piece, increasing the power of the instrument by
stronger objectives when necessary. Kellner's eye-piece
has the lens next the eye made achromatic. The peri-
scopic eye-piece of some of the German opticians has both
lenses double convex. This gives a larger field of view
with some loss of accurate definition. For high powers,
I have used a strong meniscus in place of the lower lens
in the Huygehian eye-piece. Dr. Eoyston Pigott has
suggested improvements in eye-pieces by using an inter-
mediate Huygenian combination, reversed, between the
objective and ordinary eye-piece. This gains power, but
somewhat sacrifices definition. Still better, he has pro-
posed an aplanatic combination, consisting of a pair of
slightly overcorrected achromatic lenses, mounted mid-
way between a low eye-piece and the objective. This
has a separating adjustment so as to traverse two or
three inches. The focal length of the combination varies
from one and a half to three-fourths of an inch. The
future improvement of the microscope must be looked for
in this direction, since opticians seem to have approached
the limit of perfection in high power objectives, some of
which have been made equivalent to g'oth or TJ0th of an
inch focal length. As an amplifier, I have used a double
concave lens of an inch in diameter and a virtual focus of
one and a half inches between the object-glass and the
eye:piece. If the object-glass be a good one, this will
permit the use of a very strong eye-piece with little loss
of defining power, and greatly increase the apparent size
of the object.
THE MICROSCOPE. 27
Mechanical Arrangements. — The German and French
opticians devote their attention chiefly to the excellence
of their glasses, while the mechanical part of their instru-
ments is quite simple, not to say clumsy. They seem to
proceed on the principle that as little as possible should be
done by mechanism, which may be performed by the hand.
It is different with English and American makers, some
of whose instruments are the very perfection of mechan-
ical skill. The disparity in cost, however, for instruments
of equal optical power is quite considerable.
Certain mechanical contrivances are essential to every
good instrument. The German and French stands are
usually vertical, but it is an advantage to have one which
can be inclined in any position from vertical to horizontal.
There should be steady and accurate, coarse and fine ad-
justments for focussing ; a large and firm stage with ledge,
etc., and with traversing motions, so as to follow an object
quickly, or readily bring it into the field of view ; also a
concave and plane mirror with universal joints, capable
of being brought nearer or farther from the stage, or of
being turned aside for oblique illumination. Steadiness,
or freedom from vibration, is of the utmost importance in
the construction, since every unequal vibration will be
magnified by the optical power of the instrument.
Among so many excellent opticians it would be impos-
sible to give a complete list of names whose workmanship
is wholly reliable, yet among the foremost may be men-
tioned Tolles, of Boston ; Wales, of Fort Lee, N.J.', Gru-
now, of New York ; and Zentmayer, of Philadelphia ;
Powell & Leland, Ross and Smith, Beck & Beck, of London ;
Hartnack and Cachet, of Paris ; Merz, of Munich ; and
Gundlach, of Berlin. The optical performance of lenses
from these establishments is first class, and the mechanical
work of their various models good. The finest instru-
ments from these makers, with complete appliances, are
quite costly, except the Germans and French, whose ar-
23 THE MICROSCOPIST.
rangements, as we have said, are more simple. Cheaper
instruments, however, are made by English and American
opticians, some of which are very fine.
Opticians divide microscopes into various classes, ac-
cording to the perfection of their workmanship or the
accessories supplied. The best first-class instruments have
Fio. 5. f
Wenham's Prism for the Binocular Microscope.
a great variety of objectives and eye-glasses, mechanical
stage with rack- work ; a sub-stage with rack for carrying
various illuminators : a stand of most solid construction ;
and every variety of apparatus to suit the want or wish
of the observer. They are great luxuries, although not
essential to perfect microscopic work. The second class,
or students' microscopes, have less expensive stands, but
equal optical powers, with first-class instruments. The
FIG. 6.
Collins's Harley Binocular Microscope.
30 THE MICROSCOPIST.
third or fourth classes of instruments are intended for
popular and educational use, and are fitted not only with
stands of more simple workmanship, but with cheaper
lenses, although often very good. Some French achro-
matic objectives, adapted to this class, are suitable for all
but the very finest work.
Binocular Microscopes. — The principle of the stereoscope
has been applied to the microscope, so as to permit the
use of both eyes. The use of such an instrument with
low or medium powers is very satisfactory, but is less
available with objectives stronger than one-half inch focus.
There are two ways of accomplishing a stereoscopic effect
in the microscope. The first and most common is by
means of Wenham's prism (Fig. 5), placed above the ob-
jective, and made to slide so as to transform the binocular
into a monocular microscope.
The second mode is to place an arrangement of prisms
in the eye-piece, so as to refract one-half the image to the
right and the other half to the left, which are viewed by
the corresponding eyes. In either construction there is a
provision made for the variable distance between the eyes
of different observers. In the frontispiece is a representa-
tion of Zentmayer's grand American microscope, which
will afford a good idea of the external appearance of a
first-class binocular microscope. Students' and third-class
microscopes, as before said, are less complicated and of
more moderate cost. The mechanical and optical per-
formance of Zentmayer's large instrument leaves scarcely
anything to be desired. Instead of the more expensive
rack-work stage, a simple form, originally invented by Dr.
Keen, of Philadelphia, and copied by Nachet and others,
is often employed. It consists of a rotating glass disk, to
which is attached a spring, or a V-shaped pair of springs,
armed with ivory knobs, which press upon a glass plate
in the object-carrier. The motion is exceedingly smooth
and effective.
FIG. 7.
Beck s Large Compound Microscope.
FIG. 8. FIG. 9.
Hartnack's Small Model Microscope.
Nachet's Inverted Microscope.
32 THE MICROSCOPIST.
Fig. 6 shows Collins's Harley binocular microscope, a
good second class instrument.
Fig. 7 represents Beck's large compound miscroscope
(monocular) ; and Fig. 8, Hartnack's small model micro-
scope, with the body made to incline.
Fig. 9, Cachet's inverted microscope, invented by Dr.
Lawrence Smith for chemical investigations.
CHAPTER III.
MICROSCOPIC ACCESSORIES.
IN addition to the object-glasses, eye-glasses, mirror,
and mechanical arrangement of the microscope, to which
reference was made in the last chapter, several accessory
instruments will be useful and even necessary for certain
investigations.
The. Diaphragm, for cutting off extraneous light when
viewing transparent objects, is generally needed. In some
German instruments it consists of a cylinder or tube, whose
upper end is fitted with a series of disks having central
openings of different sizes. The disk can be adjusted to
variable distances from the object on the stage so as to
vary its effects. English and American opticians prefer
the rotary diaphragm, which is of circular form, perforated
with holes of different sizes, and made to revolve under
the stage. The gradual reduction of light can be accom-
plished by the cylinder diaphragm, since when it is pushed
up so as to be near the stage it cuts off only a small part
of the cone of rays sent upwards by the concave mirror,
but, when drawn downwards, it cuts off more.
Collins's Graduating Diaphragm, which is made with
four shutters, moving simultaneously by acting on a lever
MICROSCOPIC ACCESSORIES. §3
handle, so as to narrow the aperture, accomplishes the
end most perfectly. (Fig 10.)
FIG. 10.
Collins's Now (jrr dilating Diaphragm.
Beck's Iris Diaphragm, is a further improvement of this
sort.
Condensers. — The loss of light resulting from the em-
ployment of high powers has led to several plans for con-
densing light upon the object. Sometimes a plano-convex
lens, or combination of lenses, is made to slide up and
down under the stage. A Kellner's eyepiece, or some
FIG. 11.
Smith and Beck's Achromatic Condenser.
similar arrangement, especially if fitted with a special
diaphragm, containing slits and holes, some of the latter
having central stops, is of very great use. First-class in-
struments are fitted up with achromatic condensers (Fig.
11), carrying revolving diaphragms, some of whose aper-
34 THE MICROSCOPIST.
tures are more or less occupied by stops, or solid disks, so
as to leave but a ring of space for light to pass through.
The effect of these annular diaphragms is similar to an
apparatus for oblique illumination.
The Webster condenser is similar in its optical parts to
the Kellner eye-piece, and is provided with a diaphragm
plate, with stops for oblique illumination, as well as a
FIG. 12.
Webster's Condenser, with Graduating Diaphragms.
graduating diaphragm for the regulation of the central
aperture. This is a most useful accessory. (Fig. 12.)
Oblique Illuminators — Certain fine markings on trans-
parent objects can scarcely be made out by central illumi-
nation, but require the rays to come from one side, so as
to throw a shadow. Sometimes this is well accomplished
by turning the mirror aside from the axis of the micro-
scope, and sometimes by the use of one of the condensers
referred to above. AmicCs prism, which has both plane
and lenticular surfaces, is sometimes used on one side and
under the stage, in lieu of the mirror. For obtaining
very oblique pencils of light the double hemispherical con-
denser of Mr. Reade has been invented. It is a hemi-
spherical lens of about one and a half inch diameter, with
its flat side next the object, surmounted by a smaller lens
of the same form, the flat side of which is covered with a
thin diaphragm, having an aperture or apertures close to
MICROSCOPIC ACCESSORIES.
35
its margin. These apertures may be Y-shaped, extending
to about a quarter of an inch from the centre.
If the microscope has a mechanical stage, with rack-
work, or is otherwise too thick to permit the mirror to
be turned aside for very oblique illumination, Nachefs
prism will prove of service. I have also contrived a useful
oblique illuminator for this purpose, by cementing with
Dammar varnish a plano-convex lens on one face of a to-
tally-reflecting prism, and near the upper edge of the
other side (at 90°) an achromatic lens from a French trip-
let. The prism is made to turn on a hinge, so that an
accurate pencil of light may fall on the object at any
angle desired.
Dark-ground Illuminators. — Some beautiful effects are
produced, and the demonstration of some structures aided,
by preventing the light condensed upon the object from
entering the object-glass. In this way the object appears
FIG. 13.
FIG. 14.
Nobert's Illuminator.
Parabolic Illuminator.
self-luminous on a black ground. For low powers this
can be easily done by turning aside the concave mirror as
in oblique illumination, or by employing Nobert's illumi-
nator, which is a thick plano-convex lens, in the convex
36
THE MICROSCOPIST.
surface of which a deep concavity is made. The plane
side is next the object. This throws an oblique light all
round the object. A substitute for this, called a spot lens,
is often used, and differs only from Robert's in having a
central black stop on the plane side instead of a concavity
(Fig. 13). A still greater degree of obliquity suitable for
high powers must be sought by the use of the parabolic
illuminator (Fig. 14). This is usually a paraboloid of glass,
which reflects to a focus the rays which fall upon its inter-
nal surface, while the central rays are stopped.
Illuminators for Opaque Objects. — Ordinary daylight is
hardly sufficient for the illumination of opaque objects,
FIG. 15.
Bull's-eye Condenser.
so that microscopists resort to concentrated lamplight, etc.
Gas, paraffine, and camphene lamps, have been variously
modified for this purpose, but few are better than the Ger-
MICROSCOPIC ACCESSORIES. 37
man student's Argand lamp for petroleum or kerosene
oil, as it is called. To concentrate the light from such a
source a condensing lens is used, either attached to the
microscope or mounted on a separate stand. Sometimes
a bull's-eye condenser is used for more effective illumination
(Fig. 15). This is a large plano-convex lens of short focus,
mounted on a stand. For such a lens the position of least
spherical aberration is when its convex side is towards
parallel rays ; hence, in daylight, the plane side should be
next the object. But, if it is desired to render the diverg-
FIG. 16.
Parabolic Speculum.
ing rays of a lamp parallel, the plane side should be next
the lamp, and rather close to it. The use of this con-
denser will also commend itself, when used as last referred
to, in microscopic dissection. It will throw a bright light
from the lamp directly on the trough, watch-glass, etc., in
which the specimen is being prepared. The Lieberkuhn,
or a concave speculum attached to the object-glass, and
reflecting the light from the mirror directly upon the
object, is one of the oldest contrivances for the illumina-
tion of opaque objects ; but the most convenient instru-
ment is the parabolic speculum (Fig. 16), a side mirror with
38 THE MICROSCOPIST.
a parabolic surface attached to the objective. For high
powers, a lateral aperture above the objective has been
made to throw the light down through the object-glass
itself by means of a small reflector, as devised by Prof.
Smith, or a disk of thin glass, as in Beck's vertical illumi-
nator. This latter is attached to an adapter interposed
between the objective and the body of the microscope.
Instruments for Measuring and Drawing Objects. — Screw
micrometers are sometimes used with the microscope, as
with the telescope, for the measurement of objects ; but
the less expensive and simpler glass micrometers have
generally superseded them. The latter are of two sorts,
the stage and the ocular micrometer. The stage micrometer
is simply a glass slide, containing fine subdivisions of the
inch, line, etc., engraved by means of a diamond point.
Jn case the rulings are TJDths and j-^^fhs of an inch, it
is evident that an object may be measured by comparison
with the divisions ; yet, in practice, it is found incon-
venient to use an object with the stage micrometer in this
way, and it will be found better to combine its use with
that of the drawing apparatus, as hereafter described.
The ocular, or eye-piece micrometer, is a ruled slip of glass
in the eye-piece. Its value is a relative one, depending on
the power of the objective and the length of the micro-
scope tube. By comparing the divisions with those of the
stage micrometer their value can be readily ascertained.
Thus, if five spaces of the eye-piece micrometer cover one
space of the stage micrometer, measuring fo'oo^h of an
inch, their value will be 2j0th of an inch each.
Different standards of measurement are used in different
countries. English and American microscopists use the
inch. In France, and generally in Germany, the Paris
line or the millimetre is used. The millimetre is 0.4433 of
a Paris line and 0.4724 of an English line ( ,-^th of an
inch).
In the French system the fundamental unit is the metre,
MICROSCOPIC ACCESSORIES.
39
which is the ten-millionth part of the quadrant of the
meridian of Paris. The multiples are made by prefixing
Greek names of numbers, and the subdivisions by prefix-
ing Latin names. Thus, for decimal multiples, we have
deeo, hecto, kilo, and myrio ; and, for decimal subdivisions,
deci, centi, and milli. The following may serve for con-
verting subdivisions of the metre into English equiva-
lents :
A millimetre equals 0.03937 English inches.
A centimetre " 0.39371 "
A decimetre " 3.93708 "
One inch =2.539954 centimetres, or 25.39954 millimetres.
For drawing microscopic objects the camera lucida will
be found useful. This is a small glass prism attached to
the eye-piece. iThe microscope is inclined horizontally,
FIG. 17.
Oberhauser's Drawing Apparatus.
and the observer, looking into the prism, sees the object
directly under his eye, so that its outlines may be drawn
on a piece of paper placed on the table. Some practice,
however, is needed for satisfactory results. For the up-
right stands of German and French microscopes, the camera
lucida of Chevalier & Oberhauser is available. This is a
prism in a rectangular tube, in front of which is the eye-
piece, carrying a small glass prism (c, Fig. 17), surrounded
40 THE MICROSCOPIST.
by a black metal ring. A paper placed beneath is visible
through the opening in the ring, and the image reflected
by the prism upon it can be traced by a pencil. It is neces-
sary to regulate the light so that the point of the pencil
may be seen.
Dr. Beale has recommended, in lieu of the camera lucida,
a piece of slightly tinted plate glass (Fig. 18), placed in a
short tube over the eye-piece at an angle of 45°. This is
a cheap and effective plan. A similar purpose is served
Flo. 18. Fin. 19.
Beale's Tint-glass Camera. Scemmering's Steel Disk.
by a little steel disk, smaller than the pupil of the eye,
placed at the same angle (Fig. 19).
The most simple method of measuring objects is to
employ one of the above drawing instruments, placing
first on the microscope stage an ordinary micrometer, and
tracing its lines on the paper. Then the outline of the
object can be traced and compared with the lines. The
magnifying power of an object-glass can also be readily
found by throwing the image of the lines in a stage
micrometer upon a rule held ten inches below the eye-
piece, looking at the magnified image with one eye and
at the rule with the other. Dr Beale strongly urges
observers to delineate their own work on wood or stone,
since they can do it more exactly and truthfully than the
MICROSCOPIC ACCESSORIES. 41
most skilled artists who are unfamiliar with microscopic
manipulation.
Other accessory apparatus, such as a frog-plate, for more
readily observing the circulation in a frog's foot ; an
animalcule cage, or live box ; a compressorium, for apply-
ing pressure to an object ; fishing tubes ; watch-glasses ;
growing-slides, etc., will commend themselves on personal
inspection.
For preventing the evaporation of fluids during obser-
vation, Recklinghausen invented the moist chamber (Fig.
20), consisting of a glass ring on a slide, to which is fas-
tened a tube of thin rubber, the upper end of which is
fastened round the microscope tube with a rubber band.
Recklinghauseii's Moist ( liumbcr.
A simpler form of moist chamber may be made by a
glass ring cemented on a slide. A few drops of water
cautiously put on the inner edge of the ring with a brush,
or a little moist blotting-paper may be placed inside. The
object (as a drop of frog's blood, etc.) may then be put on
a circular thin cover, which is placed inverted on the ring.
A small drop of oil round the edge of the cover keeps it
air and water-tight.
Somewhat similar to the above is Strieker's gas chamber
(Fig. 21). On the object-slide is a ring of glass, or putty,
with its thin cover. Through this ring two glass tubes
are cemented, one of which is connected with a rubber
42
THE MICROSCOPIST.
tube for the entrance of gas, while the other serves for
its exit.
For the study of phenomena in the fluids, etc., of warm-
blooded animals, we need, in addition to the moist cham-
ber, some way of keeping the object warm. This may be
roughly done by a perforated tin or brass plate on the
stage, one end of which is wrarmed by a spirit-lamp. A
piece of cocoa butter or wax will show by its melting
when the heat is sufficient. Schultze's warm stage is a
more satisfactory and scientific instrument. It is a brass
plate to fit on the* stage, perforated for illumination, and
connected with a spirit-lamp and thermometer, so that
Fro. 21.
Strieker's Gas Chamber.
the amount of heat may be exactly regulated. Other
arrangements have been proposed to admit a current of
warm water, or for the passage of electricity through an
object while under observation, which are scarcely neces-
sary to describe.
The Polariscope. — The nature and properties of polarized
light belong rather to a treatise on optics or natural phi-
losophy than to a work like the present, yet a very brief
account may not be out of place. We premise, then, that
every ray or beam of common light is supposed to have
at least two sets of vibrations, vertical and horizontal.
As these vibrations have different properties, the ray when
MICROSCOPIC ACCESSORIES.
43
divided is said to be polarized, from a fancied resemblance
to the poles of a magnet. The division of the vibrations
may be effected (i. «?., the light may be polarized) in vari-
ous ways. For the microscope the polarizer is a Nicholas
prism, composed of a crystal of Iceland spar, which has
been divided and again cemented with Canada balsam, so
as to throw one of the doubly refracted rays aside from
the field of view (Fig. 22). Such a prism is mounted in
a short tube and attached to the under side of the stage.
In order to distinguish the effects of polarized light, an
analyzer is also needed. This usually consists of another
FIG. 22.
FIG. 23.
Nichol's Pi ism.
Polarizer and Analyzer.
similar Nichol's prism, attached either to the eye-piece or
just above the objective. The latter position gives a
larger field, but the former better definition. Fig. 23
shows the polarizer and the analyzer. The polarizer is
improved by the addition of a convex lens next the object.
Hartnack has also improved the eye-piece analyzer by
adding a graduated disk and vernier.
When the polarizer arid analyzer have been put in place,
they should be rotated until their polarizing planes are
parallel, and the mirror adjusted so as to give the most
intense light. If now the polarizing planes are placed at
right angles, by turning one of them 90°, the field is ren-
44
THE MICROSCOPIST.
dered dark, and doubly refracting bodies on the stage of
the microscope appear either illuminated or in colors. If
a polarized ray passes through a doubly refracting film,
as of selenite, it forms two distinct rays, the ordinary
and the extraordinary ray. Each of these will be of dif-
ferent colors, according to the thickness of the film. Lf
one be red, the other will be green, these colors being
complementary. By using the analyzer one of these rays
is alternately suppressed, so that on revolving the appa-
ratus the green and red rays appear to alternate at each
quarter of a circle. Films of selenite are often mounted
so as to revolve between the polarizer and the stage.
Barker's selenite stage is sometimes used for this purpose
(Fig. 24). With such a stage a set of selenites is usually
i'!. 24.
Barker's Selenite Stage.
supplied, giving the blue, purple, and red, with their com-
plementary colors, orange, yellow, and green. By this
combination all the colors of the spectrum may be ob-
tained. The selenite disks generally have engraved on
them the amount of retardation of the undulations of
white light, thus: J, f, and ». If these are placed so
that their positive axes (marked PA) coincide, they give
the sum of their combined retardations.
The Microspectroscope. ^Ordinary spectrum analysis, by
determining the number and position of certain narrow
lines in the spectra of luminous bodies, called Fraunhofer's
MICROSCOPIC ACCESSORIES.
45
enables the chemist to identify different substances.
The object of the microspectroscope is different. It en-
ables us to distinguish substances by the absence of cer-
tain rays in the spectrum, or, in other words, to judge of
substances by a scientific examination of their color. The
color of a body seen with the naked eye is the general
impression made by the transmitted light, and this may
be the same although the compound rays may differ
The Sorby-Browning Microspectroscope.
greatly, so that colors which seem absolutely alike may
be distinguished by their spectra. Many solutions are
seen to absorb different colors in very definite parts of the
spectrum, forming absorption bands or lines, varying in
width and intensity according to the strength of the so-
lution. The instrument usually employed consists of a
direct-vision spectrum apparatus attached to the eye piece
of the microscope, which shows the principal Fraunhofer
46
THE MICROSCOPIST.
lines by daylight, or a spectrum of the light transmitted
by any object in the field of view. A reflecting prism is
placed under one-half of the slit of the apparatus so as to
transmit from a side aperture a standard spectrum for
comparison. In Fig. *25, A is a brass tube carrying the
compound direct-vision system of five prisms and an
achromatic lens. This tube is moved by the milled head
FIG. 2G.
Spectroscope with Micrometer.
B, so as to bring to a focus the different parts of the
spectrum. This is important when the bands or lines to
be examined are delicate. D is the stage on which objects
for comparison are placed. The light passing through
them from the mirror i, goes through a side opening to a
reflecting prism which covers a part of a slit in the bot-
tom of the tube A. This slit is opened and shut by means
of the screws c and H. Fig. 26 shows the internal ar-
MICROSCOPIC ACCESSORIES. 47
rangement of the prisms and lens, together with a microm-
eter for measuring the position of lines or absorption
bands. To use the microspectroscope, remove the tube
A, with the prisms, and insert the tube G in the place of
the eye-piece of the microscope. With the lowest power
object-glass which is suitable, and the slit opened wide by
the screw H, the object on the stage of the microscope,
illuminated by the mirror or condenser, is brought to a
focus, the tube A replaced and adjusted for focus by the
screw B, while the slit is regulated by c and H until a well-
defined spectrum is seen. To determine the position of
the absorption lines, remove the upper cover of the tube
A and replace it with that carrying the micrometer repre-
sented in Fig. 26. The mirror illuminates a transparent
line or cross, whose image is refracted by a lens c, mov-
able by a screw B, and reflected at an angle of 45° from
the upper surface of the prisms, so as to be seen upon the
spectrum. By means of the micrometer screw M, this is
made to move across the spectrum, so that the distance
between the lines may be determined. In order to com-
part the results given by different instruments, the
observer should measure the position of the principal
Fraunhofer lines in bright daylight, and mark them on
a cardboard scale, which may be preserved for reference.
By comparing the micrometric measurement of lines in
the spectrum of any substance observed by artificial light
with such a scale, their position may readily be seen.
In using the microspectroscope some objects require a
diaphragm of small size, and others, especially with the
1J or 2-inch objective, a cap with a hole j'gth of an inch
in diameter over the end of the microscope, to prevent
extraneous light from passing through the tube.
Nose-piece. — For the purpose of facilitating observations
with objectives of different powers a revolving nose-piece
has been contrived, carrying two, three, or four objectives,
48 THE M1CROSCOPIST
which may be brought quickly into the axis of the instru-
ment.
Object-finders. — It is sometimes tedious to find a small
object on a slide, particularly with high powers, and a
number of contrivances, as Maltwood's finder, have been
proposed for this end. A very simple method, however,
may serve. Mark on the stage two crosses, one like the
sign of addition -f, and the other like the sign of multi-
plication x , and, when the object is found, mark the slide
to correspond with the marks below. If the stage be a
mechanical one it will be necessary to arrange it in the
previous position.
Microscopic Photography. — Many European experimen-
ters have succeeded in taking microscopic photographs,
but a great advance in this direction has been made under
the direction of the medical department of the United
States army at Washington. Lieutenant-Colonel Wood-
ward has succeeded in furnishing permanent records of
many details of structure, which exhibit the very perfec-
tion of art. In a work like the present a full account of
the apparatus and methods employed would be out of place.
Dr. Beale's How to Work with, the Microscope, and the re-
ports issued from the Surgeon-General's office at Wash-
ington, will give the details.
CHAPTER IV.
USE OF THE MICROSCOPE.
Care of the Instrument. — But little satisfaction will be
secured in microscopic work for any length of time with-
out scrupulous care of the lenses, etc., belonging to the
instrument, and habits of this kind should be early ac-
quired. When in frequent use the microscope should be
USE OF THE MICROSCOPE. 49
seldom packed away in its case, as a certain necessary
stiffness of motion in its various parts might thereby be
lessened. Yet it should be kept free from dust and damp.
A bell-glass cover, or glass case, or a cabinet which will
admit the reception of the instrument in a form ready for
immediate use, is desirable. Before using, the condition
of objective and eye-piece should be examined as well as
of the mirror, and dust or dampness removed. Another
examination should be made before the microscope is put
away.
Stains on the brass-work may be removed by a linen
rag, and dust on the mirror and lenses by a fine camel's-
hair brush, or very soft and clean chamois skin. Frequent
wiping will injure the polish of the lenses.
The upper surfaces of the lenses in the eye-pieces and
the mirror will need the most frequent attention The
objectives, if carefully handled and kept in their boxes
when not in use, will seldom require cleaning. If the front
of the objective becomes accidentally wet with fluid it
should be at once removed, and, when reagents are used,
great care should be taken to prevent contact with the
front of the lens.
Care of the Eyes. — Continuous observation, especially by
lamplight, and with high powers, has doubtless a ten-
dency to injure the sight. To cease work as soon as
fatigue begins is, however, a simple but certain rule for
protection. This time will vary greatly, according to the
general tone and vigor of the observer. It is also impor-
tant to use the eyes alternately if a monocular instrument
is employed, as otherwise great difference both in the
focus and in the sensitiveness of the eyes will result. The
habit of keeping the unemployed eye open is a good one,
and, though troublesome at first, is not difficult to ac-
quire. It is well to protect the eye from all extraneous
light, and to exclude every part of the object except that
which is under immediate observation. The diaphragm
4
50 THE MICROSCOPIST.
will serve this end as well as modify the quality of the
light. For very del it-ate observations a dark shade over
the stage, which may be fastened by an elastic ring to
the microscope-tube, so as to shut off extraneous light,
will be useful.
Table, etc. — The microscopist's work-table should be
large and massive, so as to be convenient and free from
vibration. Drawers for accessories and materials used in
preparing and mounting objects are also desirable, as well
as a few bell-glasses for secluding objects from dust. Re-
agents should always be removed from the table after use
and kept in another place.
Light. — Dr. Carpenter has well said, " Good daylight is
to be preferred to any other kind of light, but good lamp-
light is preferable to bad daylight." A clear blue ftky
gives light enough for low powers, but a dull white
cloudiness is better. The direct rays of the sun are too
strong, and should be modified by a white curtain, reflec-
tion from a surface of plaster of Paris, or, still better, by-
passing through a glass cell containing a solution of am-
monio-sulphate of copper.
Various kinds of lamps have been contrived for micro-
scopic use ; among the best are the German and French
" student's reading lamps," which burn coal oil or petro-
leum. It is o'ften useful to moderate such a light by the
use of a chimney of blue glass, or by a screen of blue glass
between the flame and the object. Dr. Curtis contrived
a useful apparatus, consisting of a short petroleum lamp
placed in an upright, oblong box. On one side of the box
is an opening occupied with blue glass ; on another side
the opening has ground-glass, as well as a piece colored
blue, and a plano-convex lens so placed as to condense the
light thus softened to a suitable place on the table.
As a general rule the light should come from the left
side, and that position assumed or inclination given to the
instrument which is most comfortable to the observer.
USB OF THE MICROSCOPE. 51
English and American microscopists prefer an inclined
microscope, while the German and French instruments
being usually vertical do not permit this arrangement.
Adjustment. — The details of microscopic adjustments
are only to be learned by practice, yet a few directions
may be instructive. The selection of the objectives and
eye-pieces depends on the character of the object. As a
general rule, the lowest powers which will exhibit an
object are the best. It is best to use weak eye-pieces with
the stronger objectives, yet much depends on the perfec-
tion of the glasses employed.
The focal adjustment can be made with the coarse ad-
justment or quick motion when low powers are employed ;
but for higher powers the fine adjustment screw is essen-
tial. Care must be taken not to bring the objective into
close or sudden contact with the thin glass cover over the
object, and, in changing object-glasses, the microscope
body should be raised from the stage by the coarse adjust-
ment.
The actual distance between the object and object-glass
is much less than the nominal focal length, so that the
1 inch objective has a working distance of about J an
inch, the Jth of about ^th of an inch, while shorter ob-
jectives require the object to be covered with the thinnest
glass.
Sometimes, in 'high powers, and especially with immer-
sion-lenses, an adjustment of the object-glass is necessary
in order to suit the thickness of the glass cover. With
thick covers the individual lenses must be brought nearer
to each other, and, with very thin covers, moved farther
apart.
If immersion-objectives be employed a drop of water is
placed on the glass cover with a glass rod or camel's-hair
pencil, and a second drop on the lens. The lens and object
are then approximated till the drops flow together and the
focus is adjusted. By turning the Fcrew of the objective
52 THE MICROSCOPIST.
and using the fine adjustment the best position will be
shown by the sharper and more delicate image of the
object.
For other details respecting adjustment the reader is
referred to the chapter on Microscopic Accessories.
Errors of Interpretation. — True science is hindered most
of all by speculation and false philosophy, which often
assume its garb and name, but it is also retarded by im-
perfect or false observation. It is much less easy to see
than beginners imagine, and still less easy to know what
we see. The latter sometimes requires an intellect of sur-
passing endowments. The sources of error are numerous,
but some require special caution, and to these we now
refer.
The nature of microscopic images causes error from
imperfect focal adjustment. We see distinctly only that
stratum of an object which lies directly in focus, and it is
seldom that all parts of an object can be in focus together.
Hence we only recognize at once the outline of an object,
but not its thickness, and, as the parts which are out of
focus are indistinct, we may readily fall into error. Glasses
vary much in this respect. Some have considerable pene-
trating and defining power even with moderate angular
aperture, and are better for general work than those more
perfect instruments which give paler images and only re-
veal their excellencies to the practiced microscopist.
Sometimes the focal adjustment leads to error on ac-
count of the reversal of the lights and shadows at differ-
ent distances. Thus the centres of the biconcave blood-
disks appear dark when in focus, and bright when a little
within the focus ; while the hexagonal elevations of a dia-
tom, as the Pleurosigma angulatum,&re light when in focus,
with dark partitions, and dark when just beyond the
focus. From this we gather a means of discrimination,
since a convex body appears lighter by raising the micro-
scope, and a concave by lowering it.
USB OF THE MICROSCOPE. 53
The refractive power of the object, or of the medium in
which it lies, is sometimes a source of error. Thus a
human hair was long thought to be tubular, because of
the convergence of the rays of light on its cylindrical con-
vexity. A glass cylinder in balsam appears like a flat
band, because of the nearly equal refractive powers of
object and medium. The lacunae and canaliculse of bone
were long considered solid, because of the dark appear-
ance presented on account of the divergence of the rays
passing through them. Their penetration with Canada
balsam, however, proves them to be cavities. Air-bubbles,
from refraction, present dark rings, and, if present in a
specimen, seldom fail to attract the first attention of an
inexperienced observer. The difference between oil-globules
in water and water in oil, or air-bubbles, should be early
learned, as in some organized structures oil-particles and
vacuoles (or void spaces) are often interspersed. A globule
of oil in water becomes darker as the object-glass^ is de-
pressed, and lighter when raised ; while the reverse is the
case with water in oil, since the difference of refraction
causes the oil particles to act as convex lenses, and those
of water like concave lenses.
Other errors arise from the phenomena of motion visible
under the microscope. A dry filament of cotton, or other
fabric absorbing moisture, will often oscillate and twist
in a curious way.
If alcohol and water are mixed^ the particles suspended
acquire a rapid motion from the currents set up, which
continues till the fluids are thoroughly blended. Nearly
all substances in a state of minute division exhibit, when
suspended in fluid, a movement called the " Brownonian
motion," from Dr. Robert Brown, who first investigated
it. It is a peculiar, uninterrupted, dancing movement, the
cause of which is still unexplained. These movements,
as all others, appear more energetic wThen greatly magni-
fied by strong objectives. It requires care to discriminate
54 . THE MICROSCOPIST.
between such motions and the vital or voluntary motions
of organized bodies. •
The inflection or diffraction of light is another source
of error, since the sharpness of outline in an object is thus
impaired. The shadow of an opaque object in a divergent
pencil of light presents, not sharp, well-defined edges, but
a gradual shading off, from which it is inferred that the
rays do not pass from the edge of the object in the same
line as they come to it. This is in consequence of the
undulatory nature of light. When any system of wraves
meets with an obstacle, subsidiary systems of waves will
be formed round the edge of the obstacle and be propagated
simultaneously with the original undulations. For a cer-
tain space around the lines in which the rays, grazing the
edge of the opaque body,, would have proceeded, the two
systems of undulation will intersect and produce the phe-
nomena of interference. If the opaque body be very small,
and the distance from the luminous point proportionally
large, the two pencils formed by inflection will intersect,
and all the phenomena of interference will become evident.
Thus, if the light be homogeneous, a bright line of light
will be formed under the centre of the opaque object, out-
side of which will be dark lines, and then bright and dark
lines alternately. If the light be compound solar light, a
series of colored fringes will be formed. In addition to
the results of inflection, oblique illumination at certain
angles produces a double image, or a kind of overlying
shadow, sometimes called the "diffraction spectrum,"
although due to a different cause. No rules can be given
for avoiding errors from these optical appearances, but
practice will enable one to overcome them, as it were,
instinctively.
Testing the Microscope. — The defining power of an in-
strument depends on the correction of its spherical and
chromatic aberrations, and excellence may often be ob-
tained with objectives having but a moderate angle of
USE OF THE MICROSCOPE. 55
aperture. It may be known by the sharp outline given
to the image of an object, which is not much impaired by
the use of stronger eye pieces.
Resolving power is the capability an instrument has of
bringing out the fine details of a structure, and depends
mainly on the angle of aperture of the objective, or the
angle formed by the focus and the extremities of the
diameter of the lens. On this account the increase of the
angle of aperture has been a chief aim with practical
opticians.
Penetrating power is the degree of distinctness with
which the parts of an object lying a little out of focus
may be seen. Objectives which have a large angle of
aperture, and in consequence great resolving power, are
often defective in penetration, their very perfection only
permitting accurate vision of what is actually in focus.
Hence for general purposes a moderate degree of angular
aperture is desirable.
Flatness of field of view is also a necessity for accurate
observation. Many inferior microscopes hide their im-
perfection in this respect by a contracted aperture in the
eye-piece, by which, of course, only a part of the rays
transmitted by -the objective are available.
Object-glasses whose focal length is greater than half
an inch are called low powers. Medium powers range
from one-half to one-fifth of an inch focal length, and all
objectives less than one-fifth are considered high powers.
For definition with low power objectives, the pollen
grains of hollyhock, or the tongue of a fly, or a specimen
of injected animal tissue, will be a sufficient test. The
aperture should be enough to give a bright image, and
the definition sufficient for a clear image. A section of
wood, or of an echinus spine, will test the flatness of the
field.
Medium powers are seldom used with opaque objects
unless they are very small, but are most useful with
56 THE MICROSCOPIST.
properly prepared transparent objects. A good half-inch
objective should show the transverse markings between
the longitudinal ribs on the scales of the Hipparcliia
janira, butterfly (Plate I, Fig. 27), and the one-fourth or
one-fifth should exhibit markings like exclamation points
on the smaller scales of Podura plumbea (Plate I, Fig. 28)
or Lepidocyrtis.
High power objectives are chiefly used for the most
delicate and refined investigations of structure, and are
not so suitable for general work. It is with these glasses
that angular aperture is so necessary to bring out striae,
and dots, and other delicate structures, under oblique
illumination. For these glasses, the best tests are the
siliceous envelopes of diatoms, as the Pleurosigma angu-
latum, Surirella gemma, Grammataphora subtilissima ; or
the wonderful plates of glass artificially ruled by M. Ro-
bert, and known as Nobert's test.
The latter test is a series of lines in bands, the distance
between the lines decreasing in each band, until their
existence becomes a matter of faith rather than of sight,
since no glass has ever revealed the most difficult of them.
The test plate has nineteen bands, and their lines are
ruled at the following distances: Band 1, y^u^h °f a
Paris line (to an English inch as .088 to 1.000, or as 11 to
125). Band2,T5'^th. Band 3, ^th. Band 5,
Band 9, ^th. Band 13, ^th. Band 17,
Band 19, ^i^th.
It is said that Hartnack's immersion system No. 10
and oblique light has resolved the lines in the 15th band,
in which the distance of lines is about ^To^th of an inch.
The surface markings of minute diatoms are also ex-
cessively fine. Those of Pleurosigma formosiim, being from
20 to 32 in y^^th of an inch ; of P. hippocampus and P.
attenuatum about 40 ; P. angulatum 46 to 52 ; Navicula
rhomboides 60 to 111 ; and Amphipleura pelludda 120 to
130. This latter has been variously estimated at 100,000
PLATE I
FIG. 28.
Scale of Hijypnrchia Janira.
of Poilura plumbea : — A, large
O strongly marked scale; B, small scale
more faintly marked; c, portion of an
injured scale, showing the nature of the
markings.
Fio. 29.
Ple.uro.ngmn artgulntum : — A, entire frustule, as seen
under a power of 500 diam.; B, hexagonal aerolation,
as seen under a power of 1300 diam.; c, the same,
as seen under a power of 15,000 dinm.
USE OF THE MICROSCOPE.
57
to 130,000 in an inch. It has been resolved by Dr. Wood-
ward with the j'gth immersion of Powell and Lealand,
using oblique sunlight through a solution of ammonio-
sulphate of copper.
The longitudinal lines (between the transverse) of the
FIG. 30.
Valve of Surirella Gemma,
a. Transverse ridges. 6. Longitudinal lines, c. The same, resolved into areolations.
/Surirella gemma are estimated at 30 to 32 in T^o^n °f a
millimetre, and the markings on Grammataphora subtilis-
sima at 32 to 34 in the same distance.
FIG. 31.
Grammataphora Subtilissima.
a. Valve. 6. Transverse lines.
J. D. Moller has produced a very excellent test-plate,
containing twenty diatoms, with descriptions, according
to their value as tests.
58 THE MICROSCOPIST.
The Pleurosigma angulatum (Plate I, Fig. 29), with suit-
able power and illumination, should show distinct hexag-
onal areolations. The Surirella gemma (Fig. 30) shows a
series of fine transverse lines across the ridges which run
from the edge to the central line. The finest of these
ridges are not always readily seen, and the transverse
ones are only to be mastered by toil and patience.
The Grammataphora subtilissima (Fig. 31) shows trans-
verse lines (or rows of dots) along the edge, and sometimes
a double series of oblique lines.
CHAPTER Y.
MODERN METHODS OF EXAMINATION.
MICROSCOPY does not limit its researches to optical
enlargement, but seeks to comprehend elementary struc-
ture, and its methods vary according to the object imme-
diately in view. It may seek merely to discern the form
or morphology of the elementary parts or their peculiar
functions. It may be concerned with inorganic forms,
normal or pathological anatomy, or with physiology.
Each department of pursuit will suggest some variation,
yet a general plan of operation is possible.
Coarse, and moderately large objects, as a small insect,
a piece of vegetable tissue, etc., may be observed by plac-
ing it in the forceps, or on the stage of the instrument,
under an objective of low power, but ordinarily a consid-
erable degree of preparation is needed in order to acquire
a true idea of structure.
Most of the tissues to be examined are in a moist con-
MODERN METHODS OF EXAMINATION. 59
dition, and many require to be dissected or preserved in
fluid. This has much to do with the appearance of the
object in the microscope. If fibres or cells are imbedded
in connective tissue or in' fluids, of which the refractive
power is the same as their own, they cannot be perceived
even with the best glasses, and artificial means must be
resorted to that they may become visible. The refractive
power of different media causes different appearances.
Thus a glass rod lying in water is easily seen, but in
Canada balsam, whose refractive power is nearly the same
as glass, it is barely seen as a flat band, and in the more
highly refractive anise oil it presents the appearance of a
cavity in the oil.
During life the cavities and fissures in animal tissues,
in consequence of the different refractive power of their
contents and the change which takes place soon after
death, exhibit a sharpness and softness of outline which
is seldom seen in preparations.
There are two methods of microscopic investigation or
of preparation preliminary to direct observation: 1. Me-
chanical, for the separation and isolation of the elemen-
tary parts. 2. Chemical, which dissolve the connecting
material, or act on it differently than on other elements.
For minute dissection a great variety of instruments
have been proposed, but by practiced hands more can be
accomplished in shorter time by simple means than with
complicated ones. Two or three scalpels, or small ana-
tomical knives, a pair of small scissors, such as is used in
operations on the eye, and fine-pointed forceps, will be
found useful. But the most serviceable instruments are
dissecting-needles, such as the microscopist may make for
himself. A common sewing-needle, with the eye end
thrust into a cedar stick aboiit three inches long and one-
fourth of an inch diameter, will answer the purpose. The
point should not project so far as to spring, and if desired,
a cutting edge can be given to it by a hone.
60
THE MICROSCOPIST.
The light should be concentrated on the work by means
of a bull's-eye condenser, and as far as possible, the dis-
section should be carried on with the unassisted eye.
Very often the work is so fine that a magnifying glass,
or simple microscope, fixed to a suitable arm, will be
needed. A large Coddington lens, an inch and a half in
diameter, such as is used frequently by miners, will be
useful. Sometimes it is necessary to resort to the dissect-
ing microscope, which is a simple lens, of greater or less
power, arranged with rack and pinion, mirror, etc.
The specimen may be dissected under water, in a glass
or porcelain dish, or a trough made of gutta-percha, etc.
Dr. Lawson's binocular dissecting microscope (Fig. 32) is
FIG. 32.
Lawson's Binocular Dissecting Microscope.
a most useful form, as both eyes may be used. Loaded
corks, with sheet lead fastened to their under surface,
may be used to pin the subject on for greater facility in
dissection. Rests, or inclined planes of wood, one on each
side of the trough, will give steadiness to the hands.
Camels'-hair pencils for the removal of dust and extrane-
MODERN METHODS OP EXAMINATION. 61
ous elements, and for spreading out thin and delicate tis-
sues or sections, are indispensable. Pipettes, or glass
tubes, one end of which can be covered with the end of
the finger, may serve to convey a drop of fluid or a small
specimen from a bottle.
Preparation of Loose Textures. — If the formed elements
of tissue do not combine in a solid mass, it is only neces-
sary to place a small quantity on a glass slide and cover
it with a plate of thin glass. If the elements are too close
for clear definition under the microscope, a drop of fluid
may be added. The nature of this fluid, however, is not
a matter of indifference. Some elements are greatly
changed by water, etc., and it becomes important to con-
sider the fluid which is most indifferent. Glycerin and
water, one part to nine of water, will serve well for most
objects. Animal tissues are often best treated with aque-
ous humor, serum, or iodized serum. A weak solution of
salt, 7.5 grains chloride of sodium to 1000 grains of dis-
tilled water, serves for many delicate structures. (See
section on Fluid Media.}
Preparation by Teasing. — A minute fragment of tissue
should be placed in a drop of fluid on a slide, and torn or
unravelled by two sharp needles. This is accomplished
more easily after maceration, and sometimes it is neces-
sary to macerate in a substance which will dissolve the
connecting material. This picking or teasing should be
slowly and accurately performed. Beginners often fail
of a good preparation by ceasing too soon, as well as by
having too large a specimen. The most delicate manipu-
lation is required to isolate nerve-cells and processes.
Preparation by Section. — A section of soft substance may
be made with a sharp knife or scalpel, or with a pair of
scissors curved on the upper side. A section cut with the
latter will taper away at the edges so as to aflbrd a view
of its structure.
62 THE MICROSCOPIST.
Valentin's double knife (Fig. 33) is used for soft tissues
where only a moderate degree of thinness is needed. The
blades should be wet, or the section made under water.
Soft substances often require hardening before sections
can be made. The most simple and best method is that
of freezing, by surrounding the specimen with a freezing
mixture, when it may be cut with a cold knife. Small
pieces of tissue may be hardened in absolute alcohol, fre-
quently renewed. Chromic acid, in solution of one-fourth
to two per cent., is often used for animal tissues, or bichro-
mate of potash of the same strength. A solution of one-
fifth to one-tenth per cent, of perosmic acid or of chloride
of palladium is also recommended.
Soft tissues often require imbedding in a concentrated
solution of gum or of wax, spermaceti, or paraffin tem-
pered with oil. In this case sections may be made readily
by means of a section-cutter. For imbedding in wax, etc.,
FIG. 33.
Valentin's Knife.
the specimen must be hardened in alcohol, then treated
with oil of cloves or turpentine, and the section should be
mounted in Canada balsam or Dammar varnish.
Sections of hard substances, or of those imbedded, are
often made by machines invented for the purpose. One
of the simplest is (Fig. 34) an upright hollow cylinder,
with a kind of piston, pushed upwards by a fine screw.
The upper end of the cylinder carrying the specimen ter-
minates in a flat table, along which a sharp knife or flat
razor is made to slide. At one side of the tube is a
binding-screw for holding the specimen steady. A sec-
MODERN METHODS OF EXAMINATION. 63
tion may be cut by such an instrument after inserting
the structure desired in a piece of carrot, etc., which may
be placed in the tube ; or the tube may be filled with wax,
etc., and the specimen imbedded. Bones, teeth, shells,
corals, minerals, etc., require to be cut with fine saws, or
a disk of thin iron on a lapidary's wheel, and filed or
ground down to the requisite thinness, then polished with
emery, rouge, etc. The green oxide of chromium has
been suggested to me as a useful polishing powder for
hard substances. For calcareous substances, files and
hones will suffice to reduce the thickness, and putty
FIG. 34.
Section-Cutter.
powder or jewellers' rouge for polishing. They should
be mounted in Canada balsam.
Staining Tissues. — Certain elements, not previously visi-
ble, can often be made evident by certain coloring matters,
by which some constituents become more quickly or more
thoroughly stained than others. The " germinal matter,"
or "bioplasm" of Dr. Beale, identical with the "proto-
plasm " or " sarcode " of other observers, may thus be dis-
tinguished from the " formed materials " or " tissue ele-
64
THE MICROSCOPIST.
ment," which are the products of its activity. Carmine,
anilin, haematoxylin, and picric acid, are used for staining
by imparting their own color to tissues ; while nitrate of
silver, chloride of gold, chloride of palladium, and peros-
mic acid stain, by their chemical action, often under the
reducing influence of light. (See Fluid Media.}
Injecting Tissues. — Injections of the vessels in animal
tissues are resorted to either to exhibit their course or the
structure of the vascular walls. For the latter purpose a
solution of nitrate of silver is commonly employed, for the
former either opaque or transparent coloring matter. (See
Fluid Media.}
The injecting syringe (Fig. 35) is made of brass or Ger-
FIG. 35.
Injecting Syringe.
man silver. One of the pipes should be inserted into the
principal vessel, as the aorta of a small animal, the um-
bilical vein of a foetus, or the artery, etc , of an organ,
and should be securely fastened by a thread. All other
open vessels should be tied. The solution of gelatin, or
other matter used, should be strained, so as to be free
from foreign particles, and should be forced into the ves-
sels with a gentle, steady pressure on the syringe.
Injections should be made soon after the death of the
animal, or else after the rigor mortis has subsided.
MODERN METHODS OF EXAMINATION. 65
Sometimes the syringe is substituted by a self-acting
apparatus, consisting of a Wolfe's bottle, containing the
fluid, which is pressed upon by a column of air from
another source, and driven through a flexible tube to the
pipe in the bloodvessel.
The older anatomists used colored plaster or wax to
demonstrate the arteries and veins, but modern histology
requires finer materials. Isinglass or gelatin, colored, and
injected warm, or a solution of colored glycerin, are now
resorted to. The former serves for opaque, and the latter
for fine, transparent injections.
The art of injecting can only be learned by practice,
yet perseverance, in despite of many failures, will insure
success.
The liver, kidney, etc., may be injected separately, and
it is often desirable to use various colors for the different
sets of vessels. After injection thin slices may be cut off
and mounted in fluid or balsam.
Preparation in Viscid Media. — Dr. Beale has proposed a
method of preparing animal and vegetable tissues for -ex-
amination with the very highest powers, which has led to
valuable results. It consists in using pure glycerin or
strong syrup, instead of watery solutions. In this way
an amount of pressure may be applied to sections, in order
to render them thin enough for examination, which would
be destructive to specimens in water, while the preserva-
tive action of the media prevents change in the structure.
It is necessary to soak the specimen some time, and the
strength of the fluid should be gradually increased until
the tissue is permeated by the strongest that can be ob-
tained. Dr. Beale has found that minute dissection is
much more readily performed in such fluids, and that
even very hard textures, as bone and teeth, may be softened
by them, especially if acetic acid is added, so as to permit
thin sections to be made with a knife. He recommends
66 THE MICROSCOPIST.
vessels to be first injected, as with fine, transparent blue,
and the germinal matter to be stained with carmine. A
few drops of a solution of chromic acid, or bichromate of
potash, so as to impart to the glycerin a pale straw color,
serves to harden even the finest nerve-structures. Acetic
acid, and other reagents also, are much more satisfactorily
used with glycerin than with water. If syrup is used,
camphor, carbolic acid, etc , must be employed to prevent
the growth of fungi, but pure glycerin is free from this
inconvenience.
A great advantage of this mode of investigation con-
sists in the fact that a specimen thus prepared is already
mounted, and needs but a proper cement to the glass cover
and a finish to the slide, when it is ready for the cabinet.
FLUID MEDIA.
1. INDIFFERENT FLUIDS.
The vitreous humor, amniotic liquor, serum, etc., which
form the usual fluids termed indifferent, always contain
what Prof. Graham designated colloid and crystalloid
substances. In 1 000 parts there are about 4 parts of col-
loid (albumen) and 7.5 of crystalloid substance (chloride
of sodium).
The iodine serum of Schultze consists of the amniotic
fluid of the embryo of a ruminant, to which about 6 drops
of tincture of iodine to the ounce is added A small piece
of camphor will preserve this from decomposition a long
time. A substitute for this is composed of 1 ounce of
white of egg, 9 ounces of water, 2 scruples chloride of
sodium, with the corresponding quantity of tincture of
iodine.
MODERN METHODS OF EXAMINATION. 6?
'&J..
, /r
2. CHEMICAL REAGENTS'.
The greatest care should be used with these, that the
instrument and glasses maybe preserved. A small drop,
applied by a glass rod drawn out to a point to the edge of
the glass cover, will suffice in most cases,
Sulphuric Add. — Concentrated is used to isolate the
cells of horny structures, as hair, nails, etc. Dilute (1 part
to 2-3 of water) gives to cellulose, previously dyed with
iodine, a blue or purple color, and, when mixed with
sugar, a rose-red to nitrogenous substances and bile. 0.1
to 1000 of water, at a temperature of 35-40° C., resolves*
connective tissue into gelatin and dissolves it, so as to be
useful in isolating muscular fibres.
Nitric Acid. — Diluted with 4 or 5 parts water, separates
the elementary parts of many vegetable and animal tis-
sues when they are boiled or macerated in it. With chlo-
rate of potash it is still more energetic, but caution is
needed in its, use.
Muriatic Acid, Strong. — Used for dissolving intercellular
substance, as in the tubes of the kidney, etc. Dilute for
dissolving calcareous matter.
Chromic acid, J to 2 per cent, solution for hardening
nerves, brain, etc.
Oxalic acid, to 15 parts water, causes connective tissue
to swell and become transparent, while albuminoid ele-
ments are hardened. Preserves well delicate substances,
as rods of retina, etc.
Acetic acid makes nuclei visible and connective tissue
transparent, so as to exhibit muscles, nerves, etc., other-
wise invisible.
Iodine (1 grain of iodine, 3 grains iodide of potassium,
1 ounce of water) turns starch blue and cellulose brown.
Caustic potash or soda renders many structures trans-
parent.
68 THE MICROSCOPIST.
Lime-water or baryta-water is used for investigating con-
nective structures, especially tendon, as maceration en-
ables the needle to divide its fibrilla.
Chloride of Sodium. — Solutions of this salt for indifferent
media should always have some colloid, as albumen or
gum-arabic added (7.5 grains in 1000 grains of water for
delicate structures).
Bichromate of potash is used in stronger solution for the
same purposes as chromic acid.
Miiller *s eye-fluid for hardening the retina, and preserv-
ing delicate embryos, etc., consists of bichromate of potass,,
2 grammes ; sulphate of soda, 1 gramme ; distilled water,
100 grammes.
Alcohol dissolves resins and many vegetable coloring
matters ; renders most vegetable preparations more trans-
parent, and albuminous animal tissues more opaque.
Acetic acid and alcohol, 1 part of each to 2 of water,
renders connective tissue transparent, and albuminoid tis-
sue prominent. The proportions can be varied.
Alcohol and soda (8-10 drops of strong solution of caustic
soda to each ounce) renders many tissues very hard and
transparent. Beale recommends it for embryonic struc-
tures.
Ether dissolves resins, oils, and fat.
Turpentine renders dried animal sections transparent.
Oil of cloves acts as turpentine.
Solution of chloride of zinc, iodine, and iodide of potassium,
is recommended by Schacht as a substitute for iodine and
sulphuric acid to color vegetable cells, etc. Zinc is dis-
solved in hydrochloric acid, and the solution is evaporated
to syrupy consistence in contact with metallic zinc. This
is saturated with iodide of potassium, iodine added, and
the solution diluted with water. Wood cells, after boiling
in caustic potash, are stained blue by it.
Boracic acid, used by Prof. Brucke to separate the ele-
ments of red blood-corpuscles.
MODERN METHODS OF EXAMINATION. 69
3. STAINING FLUIDS.
Thiersch's Carmine Fluids.
a. RED FLUID.
1. Carmine, . . . . . . 1 part.
Caustic ammonia, . ••*> "... '.,'-... 1 "
Distilled water, . . . . . 3 parts. Filter.
2. Oxalic acid, . . . .• . . 1 part.
Distilled water, . * » -v . . 22 parts.
1 part of carmine solution to 8 parts of the acid solution, add 12 parts
absolute alcohol. Filter. After staining wash in 80 per cent, alcohol.
b. LILAC FLUID.
Borax, , • ...-.. • • • • 4 parts.
Distilled water, . . . . * . . 56 "
Dissolve and add,
Carmine, . . . . . '.-.'• 1 part.
Mix with twice the volume of absolute alcohol and filter.
Beetle's Carmine Fluids.
Carmine, . . . . . .10 grains.
Strong liquor ammonia, . . ' . . \ drachm.
Glycerin, .... . . * " . 2 ounces.
Distilled water, . . •' -.' v . 2 " ' i
Alcohol, . . . .. - , . \ ounce.
Dissolve the carmine in the ammonia in a test-tube by aid of heat ; boil
it and cool and add the other ingredients. Filter.
Acid Carmine Fluid. — Mix ammoniacal solution of car-
mine with acetic acid in excess and filter. This is said
to stain diffusely, but adding glycerin with muriatic acid
(1 : 200), concentrates the color in the cell-nucleus.
Anilin (or Magenta) Red Fluid.
Fuchsin (crystal), 1 centigramme.
Absolute alcohol, 20-25 drops.
Distilled water, ..... 15 cubic centim.
Anilin Blue Fluid. — Anilin blue, treated with sulphuric
acid and dissolved in water till a deep cobalt color is
obtained.
70 THE MICROSCOPIST.
Blue Fluid from Indigo Carmine.
Oxalic acid, 1 part.
Distilled water, 20-30 parts.
Indigo carmine to saturation.
Logwood Violet Fluid.
1. Haematoxylin, ...... 20 grains.
Absolute alcohol, $ ounce.
2. Solution of 2 grains of alum to 1 ounce of water.
A few drops of the first solution to a little of the second in a watch-
•glass, etc.
Picro-Carmine Fluid. — Filter a saturated solution of
picric acid, and add, drop by drop, strong ammoniacal
solution of carmine till neutralized.
Nitrate of Silver Fluid. — Fresh membranous tissues,
exposed to 0.5 to 0.2 per cent, solution of nitrate of silver,
washed and exposed to light, often show a mosaic of epi-
thelium, etc.
Osmic Acid. — y^th to 1 per cent, solution stains the
medulla of nerves, etc., black.
Chloride of Gold. — The solution should be similar to that
of nitrate of silver. Exposure to light stains the nerves,
etc., a violet or red color.
Prussian Blue. — After immersing a tissue in 0.5 to 1
per cent, solution of a protosalt of iron, dip it in a 1 per
cent, solution of ferrocyanide of potassium.
Other Staining Fluids. — Marked effects are often pro-
duced by the use of the violet, blue, and other inks in the
market. Thus I succeeded in some demonstrations of
nerve plexuses in muscle better than in any other way.
I suspect the particular ink employed contained a large
per cent, of soluble Prussian blue.
4. INJECTING FLUIDS.
For opaque injection several plans have been devised.
Resinous and gelatinous substances, variously colored, are
MODERN METHODS OP EXAMINATION. 71
most usual. Lieberkuhn used tallow, varnish, and tur-
pentine, colored with cinnabar ; and Hyrtl, whose prepa-
rations have been much admired, follows a similar plan.
He evaporates pure copal or mastic varnish to the consis-
tence of syrup, and grinds one-eighth as much cinnabar
and a little wax with it on a slab. For fine injections this
is diluted with ether.
For a bright red, the cinnabar may be mixed with a
little carmine
For a yellow color, the chromate of lead, prepared by
mixing solutions of acetate of lead (36 parts to 2 ounces
of water), and red chromate of potash (15 parts).
White may be made with zinc-white or carbonate of
lead — 4J ounces of acetate of lead in 16 ounces of water,
mixed with 3 J ounces carbonate of soda in 16 ounces.
For gelatinous injections the coloring matter is com-
bined with jelly, prepared by soaking fine gelatin in cold
water for several hours, then dissolving in a water-bath
and filtering through flannel.
By injecting gelatinous fluid solutions of various salts,
the coloring matter may be left in the vessels by double
decomposition.
A red precipitate, with iodide of potassium and bichlo-
ride of mercury.
A blue, by ferrocyanide of potassium and peroxide of
iron, etc.
Dr. Goadby's formula for a yellow color is :
Saturated solution of bichromate of potassium, . 8 ounces.
Water, 8 "
Gelatin, . . . . . . . 2 "
Saturated solution of acetate of lead, . . .8 ounces.
"Water, ........ 8 "
Gelatin, 2 "
For gelatinous injections, both the fluid and the subject
should be as warm as may consist with convenience.
Camphor also should be added to prevent mould.
72 THE MICROSCOPIST.
For transparent injections, gelatin may be used combined
with colored solutions, or still better, glycerin, which may
be used cold.
ThierscKs Blue. — Half an ounce of warm concentrated
solution (2:1) of fine gelatin is mixed with 6 cubic centim-
etres of a saturated solution of sulphate of iron. In
another vessel, 1 ounce of the gelatin solution is mixed
with 12 cubic centimetres of saturated solution of ferro-
cyanide of potassium, to which 12 cubic centimetres of
saturated solution of oxalic acid is added. When cold,
add the gelatinous solution of sulphate of iron drop by
drop, with constant stirring, to the other. Warm it,
still stirring, and filter through flannel.
Gerlach's Carmine. — Dissolve 5 grammes (77 grains) of
fine carmine in 4 grammes (70 grains) of water and J
gramme (8 drops) of liquor ammonia. Let it stand sev-
eral days (not airtight), and mix with a solution of 6
grammes of fine gelatin to 8 grammes of water, to which
a few drops of acetic acid are added.
Thiersch's Yellow. — Prepare a solution of chromate of
potash (1 : 11), and a second solution of nitrate of lead, of
same strength. To 1 part of the first add 4 parts of solu-
tion of gelatin (about 20 cubic centimetres to 80), and to
2 parts of the second add 4 parts of gelatin (40 cubic cen-
timetres to 80). Mix slowly and carefully, heat on a
water-bath, and filter through flannel.
Equal parts of Thiersch's blue and yellow carefully
mixed and filtered make, a good green.
COLD TRANSPARENT INJECTIONS.
Beale's Blue.
Glycerin, . . . . ... . .1 ounce.
Alcohol, . . ..'.,. . . . .1 "
Ferrocyanide of potassium, .... 12 grains.
Tincture of perchloride of iron, . . . 1 drachm.
Water, . . ., ». . .., ;» • • 4 ounces.
MODERN METHODS OF EXAMINATION. 73
Dissolve the ferrocyanide in 1 ounce of water and glyc-
erin, and the muriated tincture of iron in another ounce.
Add the latter very gradually to the other, shaking often ;
then gradually add the alcohol and water.
Scale's Finest Blue.
Price's glycerin, . . . . . « 2 ounces.
Tincture of perchloride of iron, . . . 10 drops.
Ferrocyanide of potassium, . . . 3 grains.
Strong hydrochloric acid, ..... 8 drops.
Water, 1 ounce.
Mix the tincture of iron with 1 ounce glycerin and the
ferrocyanide, after dissolving in a little water, with the
other ounce. Add the iron to the other solution gradu-
ally, shaking well. Lastly, add the water and hydro-
chloric acid. Sometimes about 2 drachms of alcohol are
added.
Mutter's Blue. — This is made by precipitation of soluble
Prussian blue from a concentrated solution by means of
90 per cent, alcohol.
Beetle's Carmine. — Mix 5 grains of carmine with a few
drops of water, and when well incorporated, add 5 or 6
drops of liquor ammonia. To this add J ounce of glyc-
erin, and shake well. Another J ounce of glycerin con-
taining 8 or 10 drops of acetic or hydrochloric acid is
gradually added. It is then diluted with J ounce of glyc-
erin, 2 drachms of alcohol, and 6 drachms of water.
Nitrate of Silver Injection. — For demonstrating the struc-
ture of the bloodvessels, the animal is bled, and a solution
of 0.25 to 1 per cent, of nitrate of silver, or a mixture of
gelatin with such a solution, is used.
5. PRESERVATIVE FLUIDS.
Canada Balsam. — This is perhaps the most common
medium used. When an object is not very transparent,
and drying will not injure it, balsam will do very well,
74 THE MICROSCOPIST.
but it is not adapted to moist preparations. Colonel
Woodward, of Washington, uses a solution of dried or
evaporated Canada balsam in chloroform or benzole.
Dammar Varnish. — Dr. Klein and other German his-
tologists prefers this to -Canada balsam. Dissolve J to 1
ounce of gum Dammar in 1 ounce of turpentine ; also J
to 1 ounce *of mastic in 2 ounces of chloroform. Mix and
filter.
Glycerin. — This fluid is universally useful to the micros-
copist. (See Preparation in Viscid Media, page 65.) Vege-
table and animal substances may be preserved in glycerin,
but if it is diluted, camphor or creasote must be added
to prevent confervoid growths. It is said, however, to
dissolve carbonate of lime.
Gelatin and Glycerin. — Soak gelatin in cold water till
soft, then melt in warm water, and add an equal quantity
of glycerin.
Gum and Glycerin. — Dissolve 1J grains of arsenious
acid in 1 ounce of wafer, then 1 ounce of pure gum arabic
(without heat), and add 1 ounce of glycerin.
Deane's Compound. — Soak 1 ounce of gelatin in 5 ounces
of water till soft ; add 5 ounces of honey at a boiling heat.
Boil the mixture, and when cool, add <5 drops of creasote
in J ounce of alcohol; filter through flannel. To be used
warm.
Carbolic Acid. — 1 : 100 of water is a good preservative.
Thwaite's Fluid.— To 16 parts of distilled water, add 1
part of rectified spirit and a few drops of creasote ; stir in
a little prepared chalk, and filter. Mix an equal measure
of camphor-water, and strain before using. For preserva-
tion of algse.
Solution of Naphtha and Creasote. — Mix 3 drachms of
creasote with 6 ounces of wood naphtha ; make a thick,
smooth paste with prepared chalk, and add gradually,
rubbing in a mortar, 64 ounces of water. Add a few
lumps of camphor, and let it stand several weeks before
MODERN METHODS OF EXAMINATION. 75
pouring off or filtering the clear fluid. Dr. Beale recom-
mends this highly for the preservation of dissections of
nerves and morbid specimens.
Ealfs Fluid. — As a substitute for Thwaite's fluid in
the preservation of algae. 1 grain of alum and 1 of bay
salt to 1 ounce of distilled water.
G oadln/s Solution. — Bay salt, 4 ounces; alum-, 2 ounces;
corrosive sublimate, 4 grains; boiling water, 4 pints.
This is the strength most generally useful, although it
may be made stronger or more dilute. It is a useful
fluid. If the specimen contain carbonate of lime, the
alum must be left out, and the quantity of salt may be
quadrupled.
Dr. Beale discards all solutions containing salts for
microscopic purposes, as they render the textures opaque
and granular.
Soluble Glass, or a solution of silicate of soda or potash,
or of both, has been proposed, but it is apt to render
specimens opaque.
Chloride of Calcium in saturated aqueous solution has
been much recommended, especially by botanists.
Acetate of Potash, a nearly saturated solution, is useful
for vegetable preparations and for specimens of animal
tissue which have been stained with osmic acid. The
latter do not bear glycerin.
Pacinian Fluid. — This is variously modified, but may
consist of corrosive sublimate,! part; chloride of sodium,
2 parts; glycerin, 13 parts; distilled water, 113 parts.
Sometimes acetic acid is substituted for chloride of so-
dium.
6. CEMENTS.
Gold Size is recommended by Dr. Carpenter as most
generally useful for thin covers. It is made by boiling
25 parts of linseed oil for three hours with 1 part of red
lead and J of as much umber. The fluid part is then
mixed with yellow ochre and white lead in equal parts,
76 THE MICROSCOPIST.
so as to thicken it, the whole boiled again, and the fluid
poured off for use.
BelVs Cement is said to be best for glycerin specimens.
It appears to be shellac dissolved in strong alcohol.
Brunswick Black is asphaltum dissolved in turpentine.
A little india-rubber dissolved in mineral naphtha is some-
times added.
Canada Balsam in chloroform or Dammar varnish (page
74) is often used as a cement.
Marine Glue. — This is most useful in building glass
cells, etc. It consists of equal parts of shellac and india-
rubber dissolved in mineral naphtha by means of heat.
Electrical Cement is made by melting together 5 parts
of rosin, 1 of beeswax, and 1 of red ochre. 2 parts of
Canada balsam added make it more adhesive to glass.
White, hard Varnish, or gum sandarac, dissolved in
alcohol and mixed with turpentine varnish, is sometimes
colored by lampblack, sealing-wax, etc.
White Zinc Cement. — Oxide of zinc rubbed up with
equal parts of oil of turpentine and 8 parts of solution of
gum Dammar in turpentine of a syrupy consistence, or
Canada balsam, chloroform, and oxide of zinc.
CHAPTER VI.
MOUNTING AND PRESERVING OBJECTS FOR THE MICROSCOPE.
FOR the permanent preservation of specimens, various
means are employed, according to the nature of the object
and the particular line of investigation desired. Few, if
any, objects show all their peculiarities of structure or
adaptation to function, and for scientific work it is often
MOUNTING AND PRESERVING OBJECTS. 77
necessary to have the same structure prepared in different
ways.
Opaque Objects have sometimes been attached by thick
gum to small disks of paper, etc., or to the bottom and
sides of small pill-boxes, or in cavities in slides of bone or
wood, but they are better preserved on glass slides, as
hereafter described.
The most convenient form of slide for microscopic pur-
poses is made of flattened crown or flint glass, cut into
slips of three inches by one inch, and ground at the edges.
Some preparations are mounted on smaller slips, but they
are less convenient than the above, which is regarded as
the standard size.
On such slides objects are fixed, and covered by a square
or round piece of thin glass, varying from ^th to -g^th
of an inch in thickness. Both slides and thin glass can
be procured at opticians' stores. Laminae of mica or talc
are sometimes used for lack of better material, but are too
soft. For object-glasses of the shortest focal length, how-
ever, it is necessary at times to resort to this sort of cov-
ering.
Great care should be taken to have both slide and cover
clean. With thin glass this is difficult, owing to its brit-
tleness. Practice will teach much, but for the thinnest
glass two flat pieces of wood covered with chamois leather,
between which the cover may lie flat as it is rubbed, will
be serviceable.
Very thin specimens may be mounted in balsam, glyc-
erin, etc., covered with the thin glass cover, and then
secured by a careful application of cement to the edges of
the cover. If, however, the pressure of the thin glass be
objectionable, or the object be of moderate thickness, some
sort of cell should be constructed on the slide.
The thinnest cells are made with cement, as gold size,
Brunswick black, etc., painted on with a camel's-hair pen-
cil. For preparing these with elegance, Shadbolt's turn-
78
THE MICROSCOPIST.
table has been contrived (Fig. 36). The slide is placed
between the springs, and while rotated, a ring of varnish
of suitable breadth is made on the glass.
A piece of thin glass (or even of thick glass) may be
perforated and cemented to the slide with marine glue by
FIG. 36.
Shadbolt's Turntable for making Cement-Cells.
the aid of heat; or vulcanite, lead, tin, gutta percha, etc.,
may be made into a cell in a similar way as seen in Fig.
37.
The perforation of thin glass may be easily performed
by cementing it over a hole in a brass plate, etc., with
marine glue, and punching it through with the end of a
FIG. 37.
Cell of Glass, Vulcanite, etc.
file. The edges may then be filed to the size of the hole,
and the glass removed by heating the brass. Thicker
glass may be bored with a file by moistening it with
turpentine.
Dry objects, especially if they are transparent, as dia-
toms, thin sections of bone, crystals, etc., may be attached
to the slide with Canada balsam, etc., covered with thin
MOUNTING AND PRESERVING OBJECTS. '79
glass, which should be cemented at the edges, and gummed
over all a strip of colored or lithographed paper, in which
an aperture has been made with a punch.
Mounting in Balsam or Dammar Varnish is suitable for
a very large proportion of objects. It increases the trans-
parency of many structures, filling up interstices and cavi-
ties, and giving them a smooth, beautiful appearance.
Very delicate tissues, as fine nerves, etc., are rendered in-
visible by it, and require other fluids, as glycerin.
Before mounting in balsam, the object should be thor-
oughly dry, otherwise a milky appearance will result. It
should then be placed in oil of cloves or of turpentine to
remove greasiuess and increase the transparency. A clean
slide, warmed over a spirit-lamp or on a hot plate, should
then have a little balsam placed on its centre, and the
object carefully lifted from the turpentine is put into the
balsam and covered with another drop. The slide should
then be gently warmed, and any air-bubbles pricked with
a needle-point or drawn aside. The thin glass cover should
be warmed and put on gently, in such a way as to lean
first on one edge and fall gradually to a horizontal posi-
tion. The slide may be warmed again, and the superflu-
ous balsam pressed from under the cover by the pressure
of a clean point upon it.
If the object is full of large air-spaces and is not likely
to be injured by heat, the air may be expelled by gently
boiling it in the balsam on the slide. If the object be one
which will curl up, or is otherwise injured by heat, the
air-pump must be resorted to. A cheap substitute for the
air-pump may be made by fitting a piston into a tolerably
wide glass tube closed at one end. The piston should
have a valve opening outwards. The preparation in bal-
sam may be placed at the bottom of the tube, and a few
strokes of the piston will exhaust the air.
To fill a deep cell with Canada balsam, it may be well
to fill it first with turpentine and place the specimen in
80* THE MICROSCOPIST.
it. Then pour in the balsam at one end, the slide being
inclined so that the turpentine may run out at the other.
Lay the cover on one edge of the cell and gradually lower
it till it lies flat. In this way air may be excluded.
The solution of balsam in chloroform needs no heat,
and has little liability of air-bubbles.
The excess of balsam round the edge of the glass cover
may be removed with a knife and cleaned with turpentine
or benzine, etc.
For animal tissues, the oil of cloves is sometimes used
instead of turpentine to increase the transparency, and a
wet preparation, as a stained or injected specimen, may
be mounted in balsam or Dammar by lirst placing it in
absolute alcohol to extract the water, then transferring
to oil of cloves or turpentine, and lastly, to the balsam.
In a reverse order, a specimen from balsam may be cleaned
and mounted in fluid.
Mounting in Fluid is necessary for the preservation of
the most delicate tissues and such as may be injured by
FIG. 38.
Spring Clip.
drying. Glycerin is perhaps the most generally useful
fluid. (See Preservative Fluids, page 73.)
For mounting in fluid, it is safer to have a thin cell of
varnish prepared first than to risk the running in of the
cement under the cover, as will be likely to occur other-
wise.
The air-pump is sometimes needed in mounting in fluid
to get rid of air-bubbles. A spring clip (Fig. 38) is also
MOUNTING AND PRESERVING OBJECTS. 81
a useful instrument for making moderate pressure on the
glass cover until the cement on its edge is dry. A drop-
ping-tube with a bulbous funnel, covered with thin india-
rubber, for taking up and dropping small quantities of
fluid, wrill also be of service.
Superfluous fluid may be removed from the edge of the
cover by a piece of blotting-paper, care being used not to
draw away the fluid beneath the cover.
As soon as objects are mounted, the slides should be
labelled before cementing is finished, otherwise time will
be lost in searching for a particular object among others,
or the name may be forgotten.
Boxes of wood or of pasteboard, with grooved racks at
the sides, are occasionally used for preserving a collection
of specimens. It is better, however, to have a cabinet
with drawers or trays so that the specimens may lie flat,
with their ends towards the front of the drawer. A piece
of porcelain on the end of the drawer is convenient for
the name of the class of objects contained, to be written
on with lead-pencil.
Collecting Objects. — The methods pursued by naturalists
generally will suffice for a large proportion of the objects
which are matters of microscopic inquiry, but "there are
others which, from their minuteness, require special search.
Many fresh-water species of microscopic organisms inhabit
pools, ditches, and streams. Some attach themselves to
the stems and leaves of aquatic plants, or to floating and
decaying sticks, etc. Others live in the muddy sediment
at the bottom of the water. A pond stick has been con-
trived for the collection of such organisms, consisting of
two lengths, sliding one within the other, so that it may
be used as a walking-cane. In a screw socket at one end
may be placed a curved knife for cutting portions of plants
which contain microscopic parasites; or a screw collar for
carrying a screw-topped bottle, which serves to bring up
a sample of liquid ; or it may have a ring for a muslin net.
6
82 THE MICROSCOPIST.
The net should be confined by an india-rubber band in a
groove, so as to be slipped off readily and emptied into a
bottle. The collector should have enough bottles to keep
organisms from each locality separate, and when animal-
cules are secured enough, air should be left to insure their
safety.
Marine organisms may be obtained in a similar way if
they inhabit the neighborhood of the shore, but others
can only be secured by means of the dredge or tow-net.
The latter may be of fine muslin sewn to a wire ring of
twelve inches diameter. It may be fastened with strings
to the stern of a boat, or held by a stick so as to project
from the side. For the more delicate organisms, the boat
should be rowed slowly, so that the net may move gently
through the water. Firmer structures may be obtained
by attaching a wide-mouthed bottle to the end of a net
made conical, and double, so that the inner cone may act
as a valve. The bottle may be kept from sinking by a
piece of cork. Such a net may be fixed to the stern of a
vessel, and drawn up from time to time for examination.
Minute organisms may be examined on the spot by
fishing them out of the bottle with a pipette, or small
glass tube, and placing them on a slide. A Goddington
or other pocket lens will suffice to show which are desir-
able for preservation.
Many of the lower animals and plants may be kept
alive in glass jars for some time. Frogs, etc., may be
kept under wire covers with a large piece of moist sponge.
Aquaria of various sorts may be procured and stocked
at small expense, and will afford a constant source of in-
struction. For fresh- water aquaria the bottom of the jar,
etc , should be covered with rich black earth, made into
a paste, and this should be surmounted with a layer of
fine washed sand. Roots of Valisneria, Anacharis, or
Char a may then be planted in the earth and the vessel
filled with water. As soon as the water is clear, put a
MOUNTING AND PRESERVING OBJECTS. 83
few fresh-water molluscs in to keep clown the growth of
confervse, especially such as feed on decayed vegetable1
matter, as Planorbis carinatus, Paludina vivipara, or Am-
phibia glutinosa. When bubbles of oxygen gas u