NEEDLES.--These are very useful and should be firmly fixed in handles

as recommended in Chapter I. It is convenient to have them of various

lengths and thicknesses. If curved by heating and bending to any

required shape they may be re-hardened by putting them whilst hot into

cold water. Dr. Carpenter also makes edged instruments by rubbing down

needles upon a hone. They are more pleasant to work with when _short_,

as the spring they have whilst _long_ robs them of much of their

firmness.

A _glass syringe_ is also useful in many operations, serving not

only to cleanse the objects but to add or withdraw liquids from the

_dissecting-trough_. This trough will now be described, as many

substances are so changed by becoming dry that it is impossible to

dissect them unless they are immersed in water during the operation.

If the object is opaque and must be worked by reflected light, a small

square trough may be made to the required size of gutta-percha, which

substance will not injure the edge of the knives, &c.; but where

transparency is necessary, a piece of thin plate-glass must be taken,

and by the aid of marine glue (as explained in Chapter IV.) the sides

affixed of the required depth. As pins, &c., cannot be used with the

glass troughs and the substance must be kept extended, a thin sheet of

cork loaded with lead in order to keep it under water may be used; but

this, of course, renders the bottom opaque. When working with many thin

substances, a plate of glass three or four inches long and two wide

will serve every purpose, and be more pleasant to use than the trough.

A drop or two of water will be as much liquid as is needed, and this

will lie very well upon the flat surface. As these are the principal

apparatus and arrangements which are requisite in dissection, the

method of proceeding in a few cases may now be noticed.

VEGETABLES.--The dissection of vegetable matter is much less

complicated than that of animal; maceration in water being a great

assistant, and in many cases removing all necessity for the use of the

knife. This maceration may be assisted by needles, and portions of

the matter which are not required may be removed by them. When, for

instance, the spiral vessels which are found in rhubarb are wanted,

some parts containing these are chosen and left in a small quantity of

water until the mass becomes soft, and this is more quickly effected

when the water is not changed. The mass must be then placed upon a

glass plate when practicable, or in the trough when large, and with

the aid of two needles the matter may be removed from the spiral

vessels, which are plainly seen with a comparatively low power; and by

conveying these to a clean slip of glass, repeating the process, and

at last washing well, good specimens may be procured. Most of these

should be mounted in some of the preservative liquids in the manner

described in Chapter IV. Many, however, may be dried on the slide,

immersed in turpentine, and then mounted in balsam; but liquid is

preferable, as it best preserves the natural appearance. Certain kinds

of vegetables require a different treatment to separate these spiral

vessels. Asparagus is composed of very hard vegetable matter, and some

have recommended the stems to be first boiled, which will soften them

to such a degree that they may easily be separated. Dilute acids are

also occasionally used to effect this; and in some instances to obtain

the _raphides_ caustic potash may be employed; but after _any_ of these

agents have been made use of, the objects must be thoroughly cleansed

with water, else the dissecting instruments (and perhaps the cell) will

be injured by the action of the remaining portion of the softening

agent.

For the dissection of _animal tissues_ it is necessary that the

instruments be in the best order as to sharpness, &c.; and as the rules

to be observed must necessarily be somewhat alike in many instances,

the treatment required by some of the objects most frequently mounted

will now be described. We may here remark that _cartilage_ can be best

examined by taking sections, which will show the arrangement of the

cells very perfectly. This, however, is plainly seen in the mouse’s

ear without any section being necessary. Glycerine, the preservative

liquids before mentioned, and Canada balsam are all used to mount it;

but perhaps the first named may be preferred in many cases.

MUSCLE.--This is what is commonly called the “flesh” of animals. If a

piece be laid upon the slide under the microscope, bundles of “fibres”

will be perceived, which with needles and a little patience may be

separated into portions, some of these being “striated,” or marked

with alternate spaces of dark and light. Some of the _non-striated_

or _smooth_ class of muscle, such as is found in intestines, may be

prepared for the microscope by immersing for a day or two in nitric

acid diluted with three or four parts of water, and then separating

with needles and mounting as soon as possible. Sometimes _boiling_ is

resorted to to facilitate the separation, and occasions little or no

alteration in the material. Specimens are often taken from the _frog_

and the _pig_, as being amongst the best, _Goadby’s solution_ being

generally used in mounting them. The muscle of insects also shows the

striæ very perfectly.

NERVE-TISSUE.--This is seldom mounted; as Dr. Carpenter observes, “no

method of preserving the nerve-tissue has been devised which makes it

worth while to mount preparations for the sake of displaying its minute

characters,” but we will mention a few particulars to be observed in

its treatment. The nerve should be taken from the animal as soon as

possible after death, and laid upon a glass slide, with a drop or two

of serum if possible. The needles may be used to clean it, but extreme

delicacy is necessary. It will be found that the nerve is tubular

and filled with a substance which is readily ejected by very slight

pressure. When the nerve is submitted to the action of acetic acid,

the outer covering, which is very thin, is considerably contracted,

whilst the inner tube is left projecting; and thus is most distinctly

shown the nature of the arrangement. Mr. Lockhart Clarke, who has made

great researches into the structure of the spinal cord, gives a part

of his experience as follows:--He takes a perfectly fresh spinal cord

and submits it to the action of strong spirits of wine. This gives the

substance such a degree of hardness that thin sections may be readily

cut from it, which should be placed upon a glass in a liquid consisting

of three parts of spirit and one of acetic acid, which renders them

very distinct. To mount these sections, they must now be steeped in

pure spirit for two hours and afterwards in oil of turpentine, and

lastly must be mounted in Canada balsam.

TRACHEÆ OF INSECTS, &C.--The nature of these was described in Chapter

III., but the method of procuring them was not explained, as this

clearly belongs to “dissection.” The larger tubes are readily separated

by placing the insect in water, and fixing as firmly as possible,

when the body must be opened and the viscera removed. The tracheæ may

then be cleaned by the aid of a camel-hair pencil, and floated upon a

glass, where they must first be allowed to dry, and then be mounted

in balsam. Mr. Quekett gives the following method of removing the

tracheæ from the larva of an insect:--“Make a small opening in its

body, and then place it in strong acetic acid. This will soften or

decompose all the viscera, and the tracheæ may then be well washed with

the syringe, and removed from the body with the greatest facility,

by cutting away the connections of the main tubes with the spiracles

by means of fine-pointed scissors. In order to get them upon the

slide, it must be put into the fluid, and the tracheæ floated upon

it; after which they may be laid out in their proper position, then

dried and mounted in balsam.” If we wish them to bear their _natural_

appearance, they must be mounted in a cell with Goadby’s fluid; but

the structure is _sometimes_ well shown in specimens mounted dry. As

before mentioned, these tracheæ terminate on the outside in openings

termed spiracles, which are round, oblong, and of various shapes. Over

these are generally a quantity of minute hairs, forming a guard against

the entrance of dust, &c. The forms of these are seldom alike in two

different kinds of insects, so that there is here a wide field for the

student. The dissection, moreover, is very easy, as they may be cut

from the body with a sharp knife or scissors, and mounted in balsam or

fluid. Many of the larvæ afford good specimens, as do also some of the

common Coleopterous insects.

TONGUES, OR PALATES, OF MOLLUSCS.--Of the nature of these, Dr.

Carpenter gives the following description:--“The organ which is

commonly known under this designation is one of a very singular nature;

and we should be altogether wrong in conceiving of it as having any

likeness to that on which our ordinary ideas of such an organ are

founded. For, instead of being a projecting body, lying in the cavity

of the mouth, it is a tube that passes backwards and downwards beneath

the mouth, its higher end being closed, whilst in front it opens

obliquely upon the floor of the mouth, being, as it were, slit up and

spread out so as to form a nearly flat surface. On the interior of

the tube, as well as on the flat expansion of it, we find numerous

transverse rows of minute teeth, which are set upon flattened plates;

each principal tooth sometimes having a basal plate of its own, whilst

in other instances one plate carries several teeth.” These palates, or

tongues, differ much amongst the Gasteropods in form and size, some

of them being comparatively of an immense length. Many are amongst

the most beautiful objects when examined with polarized light. They

must, however, be procured by dissection, which is usually performed

as follows:--The animal is placed on the cork in the dissecting-trough

before mentioned, and the head and forepart cut open, spread out,

and firmly pinned down. With the aid of fine scissors or knife, the

tongue must be then detached from its fastenings, and placed in water

for a day or two, when all foreign matter may with a little care be

removed. In what way it should be mounted will depend on the purpose

for which it is intended. If for examination as an ordinary object, it

may be laid upon the slide and allowed to dry, which arrangement will

show the teeth very well. If we wish to see it as it is naturally, it

must be mounted in a cell with Goadby’s fluid; but if it is wanted as

a polarizing object, it must be floated upon a slide, allowed to dry

thoroughly, and then Canada balsam added in the usual manner.

In the stomach, also, of some of these molluscs teeth are found, which

are very interesting objects to examine, and must be dissected out in

the same manner as the “tongues.”

Since writing the above. Dr. Alcock (whose very beautiful specimens

prove him to be a great authority in this branch) has published some

of his experience in the second volume of the third series of “Memoirs

of the Literary and Philosophical Society of Manchester.” By his

permission I make the following extract:--

“This closes my present communication on the tongues of mollusca; but

as some members may possibly feel inclined to enter upon the inquiry

themselves, I think it will not be amiss to add a few remarks on the

manner in which they are to be obtained.

“First, as to the kinds best worth the trouble of preparation. Whelks,

Limpets, and Trochuses should be taken first. Land and fresh-water

snails can scarcely be recommended, except as a special study,--their

tongues being rather more difficult to find, and the teeth so small

that they require a high power to show them properly. It would appear,

from Spallanzani’s description of the anatomy of the head of the snail,

that even he did not make out this part, although, in his curious

observations on the reproduction of lost parts, he must have carefully

dissected more snails than any other man.

“As to preserving the animals till wanted, they should simply be

dropped alive into glycerine or alcohol. Glycerine is perhaps best

where only the tongues are wanted; but it leaves the animals very soft;

and as it does not harden their mucus at all, they are very slippery

and difficult to work upon when so preserved.

“Then as to the apparatus required for dissection. In the first

place, all the work is to be done under water, and a common saucer is

generally the most convenient vessel to use. No kind of fastening down

or pinning out of the animal is needed; and, in fact, it is much better

to have it quite free, that you may turn it about any way you wish.

The necessary instruments are a needle-point, a pair of fine-pointed

scissors, and small forceps; the forceps should have their points

slightly turned in towards each other.

“A word or two on the lingual apparatus generally, and on its special

characters in a few different animals, will conclude what I have to say.

“The mode of using the tongue can be easily seen in any of the common

water-snails, when they are crawling on the glass sides of an aquarium;

it may then be observed that from between the fleshy lips a thick

mass is protruded, with a motion forwards and upwards, and afterwards

withdrawn, these movements being almost continually repeated. The

action has the appearance of licking; but when the light falls suitably

on the protruded structure, it is seen to be armed with a number of

bright points, which are the lingual teeth, so arranged as to give the

organ the character and action of a rasp.

“If you proceed to dissection, and open the head of one of these

mollusca (say, for instance, a common limpet), you will find the cavity

of the mouth almost filled with the thick fleshy mass, the front of

which is protruded in the act of feeding; and on its upper surface,

extending along the middle line from back to front, is seen the strong

membranous band upon which the teeth are set. The mass itself consists

of a cartilaginous frame, surrounded by strong muscles; and these

structures constitute the whole of the active part of the lingual

apparatus....

“But the peculiarity of the toothed membrane, which makes its name of

‘_ribbon_’ so appropriate, is, that there is always a considerable

length of it behind the mouth, perfectly formed, and ready to come

forward and supply the place of that at the front, which is continually

wearing away by use.

“In the limpet this reserve-ribbon is of great length, being nearly

twice as long as the body, and the whole of it is exposed to view on

simply removing the foot of the animal; nothing, then, can be easier

than to extract the tongue of the common limpet. But, unfortunately,

what you find in one kind of mollusc is not at all what you find in

another. In the Acmæas, for instance, which are very closely related

to the limpets, and have shells which cannot be distinguished, the

reserve portion of the ribbon has to be dug out from the substance

of the liver, in which it is imbedded, that organ being, as it were,

stitched completely through by a long loop of it.... It might be

thought a comfortable reflection that, at all events, one end of the

ribbon can always be found in the mouth; but in many cases this is

about the worst place to look for it. Perhaps it may appear strange

that in some of the smaller species, with a retractile trunk, a

beginner may very likely fail altogether in his attempt to find the

mouth; if, however, the skin of the back is removed, commencing just

behind the tentacles, there will be very little difficulty in making

out the trunk, which either contains the whole of the ribbon, as in

the whelk, or the front part of it, as in _Purpura_ and _Murex_, where

a free coil is also seen to hang from its hinder extremity.... In the

periwinkles the same plan of proceeding, by at once opening the back of

the animal, is best; and on doing so, the long ribbon, coiled up like a

watch-spring, cannot fail to be found.

“In the Trochuses, and indeed in all the _Scutibranchiata_, one point

of the scissors should be introduced into the mouth of the animal, and

an incision made directly backwards in the middle line above to some

distance behind the tentacles; the tongue is then immediately brought

into view, lying along the floor of the mouth.”

Dr. Alcock’s method of dissection will be found to differ in some

degree from the general rules before given; and when the tongue is

dissected out he washes it for one hour (shaking it now and then) in a

weak solution of potash. After cleaning thoroughly in water, it must be

mounted by one of the methods before mentioned.

Amongst insects, especially the grasshopper tribe, are found many which

possess a gizzard, armed with strong teeth, somewhat similar to those

of the molluscs. It requires great nicety of manipulation to obtain

these for the microscope; but it would be useless to attempt any

description of the process here, as the strident can only be successful

by experience in dissecting objects less difficult to manage, and by

using the most delicate instruments.

We have now considered most of those objects which require any

_peculiar_ treatment in section-cutting, &c.; but in no branch of

microscopic manipulation is experience more necessary than in this.

CHAPTER VI.

INJECTION.

1. Injection is the filling of the arteries, veins, or other vessels

of animals with some coloured substance, in order that their natural

arrangement may be made visible. This is, of course, a delicate

operation, and needs special apparatus, which I will now attempt to

describe.

2. _Syringe._--This is usually made to contain about two ounces.

On each side of the part next to the handle is a ring, so that the

finger may be thrust through it, and the thumb may work the piston

as in an ordinary syringe. The plug of the piston must be “packed”

with wash-leather, and fit so closely as to be perfectly air-tight;

and if, when it has been used awhile, it is found that some of the

liquid escapes past the plug into the back part of the body, it must

be _repacked_, which operation will be best understood by examining

the part. These syringes are made of various sizes, but in ordinary

operations the above will be all that is needed. The _nozzle_ is about

an inch long, and polished so accurately that there is no escape when

the _pipes_ are tightly placed upon it _dry_.

3. The _pipes_ are usually about an inch long, to the end of which are

affixed thicker tubes so as to fit the nozzle, as before mentioned,

whilst a short arm projects from each side of these, so that the silk

or thread which is used to tie this artery, &c., upon the thin pipe may

be carried round these arms, and all danger of slipping off prevented.

The _pipes_ are made of different sizes, from that which will admit

only a very fine needle (and this will need now and then to be cleaned,

or to be freed from any chance obstruction), to that which will take a

large pin. These sizes must always be at hand, as the vessels of some

subjects are exceedingly minute.

4. _Stopcock._--This is a short pipe like a small _straight_ tap,

which fits accurately upon the end of the syringe like the pipes, and

also takes the pipes in the same manner. The use of this is absolutely

necessary when the object is so large that one syringe full of liquid

will not fill it. If no preventive were used, some part of the liquid

would return whilst the syringe was being replenished, but the

stopcock is then turned as in an ordinary tap, and all danger of this

effectually removed.

5. _Curved needles._--These are easily made by heating common needles

at the end where the eye is situated, and bending them with a small

pair of “pliers” into a segment of a circle half an inch in diameter.

They are, perhaps, more convenient when the bent part is thrown

slightly back where it commences. The pointed end is then thrust into

a common penholder, and the needle needs no re-tempering, as the work

for which it is wanted is simply to convey the thread or silk _under_

any artery or vessel where it would be impossible to reach with the

unassisted fingers.

6. A kind of _forceps_, commonly know by the name of “bullnose

forceps,” will be constantly required during the process of injecting.

These are short, usually very strong, but not heavy, and close very

tightly by their own spring, which may be easily overcome and so

released by the pressure of the fingers. When any vessel has not been

tied by the operator, and he finds the injected fluid escaping, one

of these “bulldogs” may be taken up and closed upon the opening. This

will cause very little interruption, and the stoppage will be almost as

effectual as if it were tied.

7. When the ordinary mode of injection is employed, it is necessary

that the preparations be kept warm during the time they are used,

otherwise the gelatine or size which they contain becomes stiff, and

will not allow of being worked by the syringe. For this purpose we

must procure small earthenware or tin pots of the size required, which

will differ according to the kind of work to be done; and to each of

these a loose lid should be adapted to protect it from dust, &c. These

pots must be allowed to stand in a tin bath of water, under which a

lamp or gas flame may be placed to keep the temperature sufficiently

high to insure the perfect fluidity of the mixture. The tin bath is,

perhaps, most convenient when made like a small shallow cistern; but

some close it on the top to place the pots upon it, and alter the shape

to their own convenience.

8. We will now inquire into some of the materials which are needed in

this operation; the first of which is _size_. This substance is often

used in the form of _glue_, but it must be of the very best and most

transparent kind. To make the liquid which is to receive the colours

for the usual mode of injecting, take of this glue seven ounces, and

pour upon it one quart of clean water; allow this to stand a few hours,

and then boil gently until it is thoroughly dissolved, stirring with a

wooden or glass rod during the process. Take all impurities from the

surface, and strain through flannel or other fine medium. The weather

affects this a little as to its stiffness when cold, but this must be

counteracted by adding a little more glue if found too liquid.

9. Instead of glue, gelatine is generally used, especially when the

work to be accomplished is of the finer kind. The proportions are very

different in this case, one ounce of gelatine to about fourteen ounces

of water being sufficient. This, like glue, must be soaked a few hours

in a small part of the cold water, the remainder being boiled and

added, when it must be stirred until dissolved. A good size may be made

by boiling clean strips of parchment for awhile, and then straining

the liquid whilst hot through flannel; but when the injections are

to be _transparent_, it is of the greatest importance that the size

be as colourless as possible. For this purpose good gelatine must be

employed, as Nelson’s or Cox’s: some persons of experience prefer the

latter.

10. _Colours._--The size-solution above mentioned will need some

colouring matter to render it visible when injected into the vessels

of any animal, and different colours are used when two or more kinds

of vessels are so treated, in order that each “set” may be easily

distinguished by sight. The proportion in which these colours are added

to the size-solution may be given as follows:--

11. For--

Red 8 parts of size-solution

(by weight) to 1 part of vermilion.

Yellow 6 ” ” 1 ” chrome yellow.

White 5 ” ” 1 ” flake-white.

Blue 3 ” ” 1 ” blue-smalt, fine.

Black 12 ” ” 1 ” lamp-black.

Whichever of these colours is made use of must be levigated in a mortar

with the addition of a very small quantity of water until every lump

of colour or foreign matter is reduced to the finest state possible,

otherwise in the process of injecting it will most likely be found

that some of the small channels have been closed and the progress

of the liquid stopped. When this fineness of particles is attained,

warmth sufficient to render the size quite fluid must be used, and the

colour added gradually, stirring all the time with a rod. It may be

here mentioned that where one colour only is required, vermilion is,

perhaps, the best; and blue is seldom used for opaque objects, as it

reflects very little more light than black.

12. When it is wished to fill the capillaries (the minute vessels

connecting the arteries with the veins), the “Micrographic Dictionary”

recommends the colouring matter to be made by double decomposition. As

a professed handbook would be, perhaps, deemed incomplete without some

directions as to the mode of getting these colours, I will here make

use of those given in that work. For red, however, vermilion, as above

stated, may be used; but it must be carefully examined by reflected

light to see whether it be free from all colourless crystals or not.

It must first be worked in a mortar, and then the whole thrown into a

quantity of water and stirred about; after leaving it not longer than

a quarter of a minute, the larger portions will settle to the bottom,

and the liquid being poured off will contain the finer powder. This may

then be dried slowly, or added to the size whilst wet in the manner

before advised.

13. _Yellow injection._--To prepare this, take--

Acetate (sugar) of lead 380 grains.

Bichromate of potash 152 ”

Size 8 ounces.

Dissolve the lead salt in the warm size, then add the bichromate of

potash finely powdered.

Some of the chromic acid remains free, and is wasted in this solution,

so the following is given:--

Acetate of lead 190 grains.

Chromate of potash (neutral) 100 ”

Size 4 ounces.

The first of these has the deepest colour, and is the most generally

used.

14. _White injection._--This is a carbonate of lead:--

Acetate of lead 190 grains.

Carbonate of potash 83 ”

Size 4 ounces.

Dissolve the acetate of lead in the warm size, and filter through

flannel; dissolve the carbonate of potash in the smallest quantity of

water, and add to the size: 143 grains of carbonate of soda may be

substituted for the carbonate of potash.

15. For blue injection, which is not, however, much used with reflected

light, as before stated, take--

Prussian blue 73 grains.

Oxalic acid 73 ”

Size 4 ounces.

The oxalic acid is first finely powdered in a mortar, the Prussian blue

and a little water added, and the whole then thoroughly mixed with the

size.

16. It may here be repeated, that it is only when the capillaries are

to be filled that there is any need to be at the trouble to prepare

the colours by this double decomposition; and, indeed, colours ground

so finely may be procured that the above instructions would have been

omitted, had it not been supposed that some students might find a

double pleasure in performing as much of the work as possible by their

own unaided labours.

17. The process of injection may now be considered; but it is

impossible for written instructions to supply the place of experience.

I will do my best, however, to set the novice at least in the right

way. There are two kinds of injection--one where the object and colours

are opaque, and consequently fit for examination by _reflected_ light

only; the other, where the vessels are filled with transparent colours,

and must be viewed by _transmitted_ light. The first of these is most

frequently employed, so we will begin with it. In the object which

is to be injected a vessel of the kind which we wish to be filled

must be found; an opening must then be made in it to allow one of the

small pipes before mentioned to be thrust some distance within it.

When this is accomplished, thread the curved needle with a piece of

silk thread, or very fine string, which some operators rub well with

beeswax. This thread must not be too thin, else there is danger of

cutting the vessel. The cord is then carried under the inserted pipe,

and the vessel bound tightly upon it, the ends being brought up round

the transverse arms, and there tied; so that all danger of accidentally

withdrawing the pipe is obviated. Care must now be used in closing

all the vessels which communicate with that where the pipe is placed

lest the injecting fluid escape; and this must be done by tieing them

with silk. Should, however, any of these be left open by accident, the

bullnose forceps must be made use of, as before recommended.

18. The part to be injected must now be immersed in warm water,

not, however, above 100° Fahrenheit, and be left until the whole is

thoroughly warmed. Whilst this is being done, the coloured size must

be made ready by the pot being placed in the tin bath of warm water,

which must be of sufficient temperature (about 110° Fahrenheit) to

keep it perfectly liquid. For the same purpose, the syringe is often

tightly covered with two or three folds of flannel; and, indeed,

there is no part of the process which requires more attention. If the

substance to be injected is too hot, it is injured; whilst, if any of

the articles are too cold, the gelatine, or size, loses a part of its

fluidity, and consequently cannot enter the minute parts. When all is

prepared, the syringe, with the stopcock attached, should be warmed,

and then filled and emptied with the injecting fluid two or three

times, care being taken that the end of the syringe be kept beneath any

bubbles which form upon the surface. The syringe may then be filled,

and closely attached to the pipe which is tied in the vessel. With a

firm and steady pressure the piston must be forced downwards, when the

substance will be perceived to swell, and the colour show itself in

places where the covering is thin. When the syringe is _almost_ emptied

of its contents, the stopcock must be turned to prevent any escape

of the injection from the subject. It must then be refilled, as in

the first instance, and the process repeated. I say _almost emptied_,

because it is well not to force the piston of the syringe quite to the

bottom, lest the small quantity of air which frequently remains be

driven into some of the vessels, and the object be injured or quite

ruined. As the injection is proceeded with, it will be found that the

force required grows greater, yet care must be taken not to use too

much, or the vessels will burst, and render all the labour fruitless.

The movement of the piston must be occasionally so slow as to be almost

imperceptible, and for this reason it is sometimes marked with lines

about one-eighth of an inch apart.

19. Of course, during the whole process the injecting fluid and subject

must be kept at a temperature high enough to allow the liquid to flow

freely; and the escape of a little of it need cause no fears to the

student, as it is almost impossible to fill any subject without some

loss. When the injected object has received sufficient fluid, it should

have a plump appearance, owing to all the vessels being well filled.

The vessel must then be tied up where the pipe was inserted, and the

whole left in cold water two or three hours, after which time it may be

mounted; but it may be well to notice a few things which the beginner

ought to know before entering into that part of the process; and he

may be here informed that it is not necessary to mount the objects

immediately, otherwise it would be impossible for one person to make

use of half of any large subject, as it would be in a state of decay

long before each part could have been examined and separated. Large

pieces should be therefore immersed in equal parts of spirits of wine

and water, or glycerine, which some think better still, and thus

preserved in bottles until time can be given to a closer examination.

20. In operating upon large subjects, entire animals, &c., the constant

pressure required by the piston of the syringe grows wearisome, besides

occupying both hands, which is sometimes inconvenient when working

without assistance. To obviate this, another way of driving the syringe

was published in the “Micrographic Dictionary” which I will quote

here:--“We have therefore contrived a very simple piece of apparatus,

which any one can prepare for himself, and which effects the object

by mechanical means. It consists of a rectangular piece of board, two

feet long and ten inches wide, to one end of which is fastened an

inclined piece of wood (equal in width to the long board, and one foot

high). The inclined portion is pierced with three holes, one above the

other, into either of which the syringe may be placed--the uppermost

being used for the larger, the lowermost for the smaller syringe; and

these holes are of such size as freely to admit the syringe covered

with flannel, but not to allow the rings to pass through them. The

lower part of the syringe is supported upon a semiannular piece of

wood, fastened to the upper end of an upright rod, which slides in

a hollow cylinder fixed at its base to a small rectangular piece of

wood; and by means of a horizontal wooden screw, the rod may be made to

support the syringe at any height required. The handle of the syringe

is let into a groove in a stout wooden rod connected by means of two

catgut strings with a smaller rod, to the middle of which is fastened

a string playing over a pulley, and at the end of which is a hook for

supporting weights, the catgut strings passing through a longitudinal

slit in the inclined piece of wood.” When in use the syringe is filled

with injecting fluid, and passed through one of the three holes which

is most suitable. The object being placed so that the pipe and syringe

can be best joined, the rod and strings are set in order, and a weight

placed on the hook. The stopcock must then be opened gradually, when

the operator will be able to judge whether the weight is a proper one

or not: if the piston is driven with any speed, there is danger of

injuring the subject, and less weight may be used; if, however, the

piston does not move, more must be added.

21. Such is the method recommended by the “Micrographic Dictionary,”

and perhaps it is as good as any mechanical plan could be; but where

the operator is willing to undergo the labour of performing all this

with the hand, he has a much better chance of succeeding, because the

pressure can be regulated so accurately, and changed so quickly when

requisite, that no mere machine can compete with it, however well

contrived.

22. When the beginner attempts to inject a subject, one of his

difficulties is finding the vessel from which to commence. Another

consists in distinguishing the arteries from the veins; but this is

partly removed by making a longitudinal incision in the vessel, and

with a blunt thick needle probing a little distance into the tube. The

artery will be found thicker in the coating than the vein, and the

difference is easily perceived by this mode of testing: the vein is

also of a bluer colour than the artery. I say above, a “longitudinal

incision” must be made: the reason for this is, the artery when cut

across contracts considerably, and is lost in the adjoining substance;

but where the opening is made _longitudinally_ all danger of this

contraction is obviated.

23. The different systems of vessels are often injected with various

colours, so that their relative positions, &c., may be shown most

clearly. In some specimens, the veins are injected with white, and the

arteries with red; in the kidney, the urinary tubes are often filled

with white, and the arteries with red. Then, again, the liver affords

tubes for three or four colours. But no written instructions on this

point can benefit the young student, and he must be content for a

while to employ himself with single colours until he has gained the

mechanical skill and the primary knowledge which are necessary before

he can make any advance.

24. We will now consider the best methods of _mounting_ injected

objects. They must always be well washed in water after they have been

kept in any preservative liquid, using a camel-hair pencil to clean the

surface if necessary. Many parts when injected are in masses, such as

the lungs, liver, &c., of animals, and consequently sections of these

must be cut. For this purpose Valentine’s knife is very convenient,

as the thickness can be regulated so easily; but where the injections

are opaque, there is no need to have the sections very thin. Some few

of this kind undergo comparatively little change in drying, so that

the section may be well washed and floated upon the glass slide in the

place desired, where it will dry perfectly and adhere to it. It must

be then moistened with turpentine and mounted in Canada balsam like

other objects. No great heat should be used with these preparations,

as it is very liable to injure them; and some of the colours seem to

suffer a slight contraction when any great degree of warmth is applied.

There are many objects, however, which must be seen in the mass to be

understood, and, indeed, lose all their form and beauty in drying, such

as certain parts of the intestines, &c. These must be mounted in fluid,

with the precautions noticed at length in Chapter IV., and for this

purpose either Goadby’s fluid, the chloride of zinc solution, or spirit

diluted with ten parts of distilled water, may be employed. It is a

good thing, when practicable, to mount similar objects on two separate

slides, using different preservative liquids, and taking the precaution

of marking each with the kind of liquid employed. This not only serves

as a guide to what is best for certain subjects, but if one is injured,

there will probably be a good specimen in the other.

25. It may be here mentioned that many are now mounting sections of

injected substances with the balsam and chloroform before mentioned,

instead of using balsam alone, and consider that the labour is much

lessened thereby.

26. A description of that mode of injection which is most generally

employed has now been given, but this is not the only method of

effecting our object. A most ingenious process was invented by M.

Doyers, requiring no artificial warmth, by which many beautiful objects

have been prepared. Make a solution of bichromate of potash, 524 grains

to a pint of water, and throw this into the vessels to be injected;

then take 1,000 grains of acetate of lead dissolved in half a pint

of water, and force this into the same vessels. A decomposition now

takes place in the vessels, and the yellow chromate of lead is formed.

In this decomposition, however, the acetate of potash also is formed

and as this salt has an injurious action upon the cells, Dr. Goadby

recommends nitrate of lead to be used, which preserves rather than

destroys them. He also advises the addition of two ounces of gelatine

dissolved in eight ounces of water, to eight ounces of the saturated

solution of each salt; but with this addition the hot-water bath would

be required to keep the injecting fluid liquid.

27. Many of these are best mounted in balsam, in the same manner as

those made in the ordinary way; whilst others are best shown when

preserved in liquids, for which purpose Goadby’s fluid may be employed.

28. This mode of making injections with chromate of lead is deemed by

many the best, especially where one colour only is employed. But it

must be allowed that there is a little more danger of failure where two

separate fluids are used for the same vessels.

29. We will now consider the best manner of making _transparent_

injections, which, for many purposes, possess an undoubted advantage

over the opaque ones. But it must be remembered that there are certain

subjects to which no transparent injection could be applied, as they

are too thick when in their natural state, and cutting would destroy

all that beauty which is shown by the different parts in their

relative adaptation. For those objects, however, which must be cut

into sections to display their wonders, or are naturally thin--such

as some of the finer tissues, livers, kidneys, &c.--transparency is a

great acquisition, and enables us to understand the arrangement of the

vessels more perfectly. Again, another advantage is the simplicity of

the process; no hot water is needed with some preparations, either for

the subject or the injecting fluid, which runs into the minute vessels

thoroughly and easily, whilst the cost is small.

30. For this kind of injection no colour is so commonly made use of

as Prussian blue. It is not a good one, as was before stated, for

any opaque object, as the light reflected from it appears almost

black; yet by transmitted light no colour is more useful, because

its distinctness is equally great by artificial light and ordinary

daylight. The method of preparing this, as given by Dr. Beale, is as

follows:--

Glycerine 1 ounce.

Wood naphtha, or pyroacetic acid 1½ drachm.

Spirits of wine 1 ounce.

Ferrocyanide (yellow prussiate) of potash 12 grains.

Tincture of sesquichloride of iron 1 drachm.

Water 4 ounces.

Dissolve the ferrocyanide of potash in one ounce of the water;

add the tincture of sesquichloride of iron to another ounce. Mix

these solutions gradually together, shaking the bottle well which

contains them--it is best to add the iron to the potash solution.

When thoroughly mixed, these solutions should produce a dark-blue

mixture, perfectly free from any perceptible masses or flocculi. Next

mix the naphtha and spirits of wine, and add the glycerine and the

remaining two ounces of water. This must now be slowly mixed with the

blue liquid, shaking the whole well in a large bottle whilst the two

come together. The tincture of sesquichloride of iron is recommended,

because it can always be obtained of a uniform strength.

31. Dr. Turnbull used a mixture slightly different from the above,

which is made with the sulphate of iron:--

Purified sulphate of iron 10 grains.

Ferrocyanide of potassium 32 grains.

Glycerine 1 ounce.

Pyroacetic acid 1½ drachm.

Alcohol 1 ounce.

Water 4 ounces.

Dissolve the sulphate of iron in one ounce of the water, gradually add

the ferrocyanide of potassium dissolved in another ounce, and proceed

as above.

32. Dr. Beale also gives us the following carmine injection to be

employed in the same way as the blue.[G] Take--

Carmine 5 grains.

Glycerine, with 8 or 10 drops of hydrochloric acid ½ ounce.

Glycerine (pure) 1 ounce.

Alcohol 2 drachms.

Water 6 drachms.

Mix the carmine with a few drops of water, and when well incorporated

add about five drops of liquor ammoniæ. To this dark-red solution about

half an ounce of the glycerine is to be added, and the whole well

shaken in a bottle. Next, very gradually pour in the acid glycerine,

frequently shaking the bottle during admixture. Test the mixture with

blue litmus-paper, and if not of a very decidedly acid reaction, a few

more drops of acid may be added to the remainder of the glycerine and

mixed as before. Lastly, mix the alcohol and water very gradually,

shaking the bottle thoroughly after adding each successive portion

till the whole is mixed. This fluid may be kept ready prepared, and

injections made very rapidly with it.

[G] When, however, it is desirable to cut very thin sections of

the injected subject, the carmine is sometimes added to a

solution of fine gelatine--gelatine one part to water eight

parts. But the warm water and mode of proceeding which

are used with the size solutions before described will be

necessary in this case also.

33. The method of making injections with these colours is the same as

with the gelatine mixtures before described, except that no heat is

required, and consequently most of the trouble removed. The bottle of

the fluid must be well shaken immediately before use; and when the

object is injected, we must allow it to remain in a cool place for a

few hours before cutting it. Thin sections of the subject may be cut

with Valentine’s knife, as before described, and are very beautiful

transparent objects. Some of the finer tissues, also, are shown much

better by this mode of injection than by the opaque, and are easily

mounted by washing in clean water when first separated, and floating

upon a slide, where they must be allowed to dry thoroughly. They may

then be immediately mounted in balsam, or kept in the dry state until

it is convenient to finish them; but in many cases this keeping, if too

much prolonged, will injure the object. If it is desired to transfer

the section to another slide, it will be necessary to wet it thoroughly

with water by the aid of a camel-hair pencil, and then gently strip it

off with the forceps. When it is wished to preserve injected subjects

in “masses,” it must be done by immersion in spirit, and the sections

may be cut at leisure. Most of these transparent objects may be mounted

in Canada balsam; but some recommend glycerine or glycerine jelly, as

allowing the use of a higher power in their examination, and preserving

them in a more natural form.

34. A few subjects may be noticed which are very beautiful when

injected, and amongst these are the eyes of many animals. They

must be injected by the artery in the back part, and when the blue

transparent liquid is employed, nothing can exceed the delicate beauty

which some of the membrane bears. It must, however, be dissected with

care, but well repays us for the trouble. Water-newts and frogs are

not difficult subjects, and in their skin and other parts are many

interesting objects. Amongst the commoner animals--rats, rabbits, cats,

&c. &c.--almost endless employment may be found, making use either of

portions or the whole animal at once. The intestines of many of these

are very beautiful. We must divide them with a pair of scissors along

the tube, and cleanse them from all the matter; the coating may then be

laid upon a slide, and any remaining impurity removed with a camel-hair

pencil and water. When dried it may be mounted in balsam, and having

been injected with the transparent blue, its minute beauty is shown

most perfectly. In injecting a sheep’s foot, which is a good object,

the liquid should be forced into it until a slight paring of the hoof

shows the colour in the fine channels there.

35. When the lungs of small animals are injected, the finest fluid must

be used, as some of the capillaries are so small that it is not an easy

matter to fill them properly. And before entering upon these subjects,

a certain proficiency in the mode of using the syringe, &c., should be

obtained by practising upon simpler parts.

36. No subjects are more difficult to inject than fish, owing to the

extreme softness of their tissues. Dr. Hogg recommends the tail of the

fish to be cut off, and the pipe to be put into the divided vessel

which lies just beneath the spinal column; by which method beautiful

injections may be made. The gills, however, are the most interesting

part as microscopic objects.

37. These instructions may seem very imperfect to those who have had

much experience in this branch; but they will remember that their

own knowledge was not gained from any written descriptions, but was

forced upon them by frequent failures, some of which probably were

very disheartening. As I before stated, it is very difficult (if not

impossible) to accomplish much without some knowledge of anatomy.

38. I may here mention that the transparent injections sent over from

the Continent are beautifully executed by Hyrtl of Vienna (who states

that the injected fluid is composed of gelatine and carmine), Dr.

Oschatz of Berlin, the Microscopic Institute of Wabern, Schaffer and

Co. of Magdeburg, and others. Some of these will bear examining with a

high power. A friend informs me that he measured a vessel in a rat’s

tongue by Hyrtl, which was 1-7200th of an inch in diameter, and had

a clear outline with quarter-inch objective. He has also made many

experiments with the same materials, but has as yet failed in producing

perfectly distinct outlines, there being a tendency of the colouring

matter (magenta, carmine. &c.) to diffuse itself through the coats of

the vessels into the surrounding tissues, although he has varied the

pressure from one half a pound to sixty pounds. He believes the vessels

are first washed out (injected with warm water and pressure applied),

then some fluid introduced which renders the arteries impervious to the

coloured fluid afterwards injected.

39. He finds that after washing out the vessels as above, the injecting

fluid is much more easily introduced. He has used a strong solution

of gallic acid previously to injecting with the colouring matter

(in one experiment only), and the result was satisfactory. He puts

the query,--Might not carbolic acid have a similar effect? He has

often used it with injections to preserve the specimens, but not in

sufficient quantity to act in the way indicated above.

Since writing the above, Mr. J. G. Dale, F.C.S., and I have made

numerous experiments with carmine injection, and have at length

been favoured with what we deem success. Some of the vessels in a

kitten lately injected do not exceed 1-2000th of an inch in diameter,

and present a clear outline with one-fifth objective. There is no

extra-vasation, neither does the colouring matter show any grain except

when a very high power is employed. The following is our process:--

Take 180 grains best carmine.

½ fluid ounce of ammonia, commercial strength, viz.,

0·92, or 15° ammonia meter.

3 or 4 ounces distilled water.

Put these into a small flask, and allow them to digest without

heat from twenty-four to thirty-six hours, or until the carmine is

dissolved. Then take a Winchester quart bottle, and with a diamond mark

the spot to which sixteen ounces of water extend. The coloured solution

must be filtered into the bottle, and to this pure water should be

added until the whole is equal to sixteen ounces.

Dissolve 600 grains potash alum in ten fluid ounces of water, and add

to this, under constant boiling, a solution of carbonate of soda until

a slight permanent precipitate is produced. Filter and add water up

to sixteen ounces. Boil and add the solution to the cold ammoniacal

solution of carmine in the Winchester quart, and shake vigorously for

a few minutes. A drop of this placed upon white filtering-paper should

show no coloured ring. If much colour is in solution the whole must

be rejected, because, although it is possible to precipitate all the

colouring matter by the addition of ammonia or alum, it is not well to

do so, as the physical condition of the precipitate is thereby altered.

Supposing the precipitation to be complete, or very nearly so, shake

vigorously for at least half an hour, and allow it to stand until quite

cold. The shaking must then be renewed for some time, and the bottle

filled up with pure water.

After allowing the precipitate to settle a day, draw off the clear

supernatant fluid with a syphon. Repeat the washing until the clear

liquid gives little or no precipitate with chloride of barium. So much

water must be left with the colour at last that it shall measure forty

fluid ounces.

For the injecting fluid take twenty-four ounces of the above coloured

liquid, and three ounces of good gelatine. Allow these to remain

together twelve hours, and then dissolve by the heat of a water bath;

after which it should be strained through fine muslin.

As this injecting fluid contains gelatine, the hot water, and other

contrivances mentioned in a former part of the chapter, will be

necessary here also, but no peculiar treatment will be required.

CHAPTER VII.

MISCELLANEOUS.

It must be evident to all readers that there are various objects

of interest to the microscopist which cannot be properly placed

amongst any of the forementioned classes, but must not be omitted in

such a guide as this professes to be. Of these may be mentioned the

circulation of the blood in various animals, the rotary motion of the

fluid in many plants, the best means of taking minute photographs, &c.

&c.

Perhaps the most interesting of these objects is the circulation of

the blood through the finer vessels of various parts of the animals

made use of for these purposes, which parts, it is evident, must be

very transparent to afford a perfect view of this phenomenon. The

web of the frog’s foot is very frequently made use of, but requires

a certain arrangement, which we will now describe. A piece of thin

wood (Dr. Carpenter recommends _cork_) is taken, about eight inches

long and three wide; about an inch from one end is cut a hole, half or

three-quarters of an inch in diameter. The body of the frog is then

placed in a wet bag, or wrapped in wet calico, whilst the hind-foot

projects; the whole is then laid upon the piece of wood so that the

foot, which is left free, may be extended over the hole. The web must

then be spread out, and secured either by threads to small pins on

the wood, or the pins must be driven through the web into the wood,

and so kept in position. A few bands of tape must be passed round the

body, the leg, and the wood, to prevent any disarrangement arising from

the animal starting, &c. Care must be taken that the tape is not too

tight, else the circulation will be very slow or altogether stopped.

The wood must now be fixed upon the stage, with the aperture under

the object-glass: this is sometimes done by simply binding it, or a

spring is fixed so as to accomplish the same object without so much

trouble. With a half-inch power the blood may now be seen to flow very

distinctly. The frog may be used for hours if care is taken to prevent

the web from becoming dry, by wetting it with a little water from time

to time. The piece of wood or cork upon which the frog is laid is

often made to give place to the “frog-plates,” supplied by opticians.

These are made of brass, somewhat resembling the piece of wood above

recommended, but each maker’s pattern differs according to his own

taste.

The tongue of the frog is also sometimes used for the purpose of

showing the circulation of the blood, which is done in the following

manner:--The body is wrapped with the calico, and made fast to the

plate as before, only the _mouth_ of the frog is brought to the

opening. The tongue is then gently drawn out of the mouth and pinned

down over the aperture, when the circulation will be well shown. But,

as Dr. Carpenter observes, the cruelty of this mode of treatment is so

repulsive that it is unjustifiable.

Tadpoles of the frog (which, of course, are only obtainable in their

season) are good subjects for showing the circulation of the blood.

They are best suited for the microscope when about one inch long. The

tadpoles of the newt and toad also are equally suitable. They may

be placed in a very shallow glass trough with a little water, and a

narrow band of linen bound lightly round in some part not required for

examination, to keep them from moving; or they may be laid upon a glass

plate with a drop or two of water, and a thin glass covering lightly

bound upon it. Dr. Carpenter, however, places them first in cold water,

gradually adding warm until the whole becomes about 100°, when the

tadpole becomes rigid, whilst the circulation is still maintained. I

have not, however, found this necessary, the thin glass accomplishing

all that is desired. The tail is generally the most transparent, and

shows the circulation best; but in some of the newt larvæ the blood may

be traced down to the very extremities if they are not _too old_. Mr.

Whitney places the tadpole upon its back, by which means the heart and

other internal arrangements may be seen.

Amongst fishes also may be found subjects for the same purpose, but

they seldom furnish as good examples as those before mentioned, because

the blood-vessels are not nearly so abundant, as in the foot of the

frog, &c. The stickleback is, however, procurable almost in any place

during the summer months, and may be laid in a shallow trough, loosely

bound down as the tadpole. The tail may be covered with a piece of thin

glass to prevent him curling it to the object-glass. The power needed

with this will be about the same as with the other subjects--viz., half

to quarter inch object-glass.

It is not absolutely necessary to go to reptiles or fishes for this

curious sight, as some other animals serve very well. In the wings of

the common bat may be found a good subject. These must be stretched out

on something resembling the frog-plate before described, when those

parts near to the bones will show the _largest_ vessels very clearly.

The ear of a young mouse is an illustration of the same phenomenon, but

it is very difficult to fix it in a good position, as these animals are

so very timid and restless.

Amongst insects also the same law may be observed, by placing them in

the “cage,” or “live-box,” so as to keep them still, but not to injure

them by too much pressure. In certain larvæ this is particularly well

shown, as that of the day-fly and plumed gnat; but in some of these

the blood is almost colourless. In the wings also of many insects this

circulation is well seen, as in those of the common housefly; but as

these parts become dry in a few days, the subject should not be more

than twenty-four hours old.

Somewhat approximating to the forementioned phenomenon, is the

“rotation” of fluid in the cells, or, as it is usually termed, the

_circulation of the sap_, of plants. This is shown in certain vegetable

growths as a constant stream of thick fluid, wherein small globules

are seen; which stream flows round the individual cells, or up the

leaf, turning at the extremity, and down again by a different but

parallel channel. There is little or no difficulty in showing this in

many plants; but some are, of course, better than others, and require

a different treatment; we will, therefore, notice a few of these.

Perhaps the best of all is the _Vallisneria spiralis_, which is an

aquatic plant, frequently grown in, but not really belonging to, this

country. As it somewhat resembles grass, the leaf is not used in its

natural state, but a thin section cut lengthwise with a razor or other

sharp instrument--this section, however, is much better when the outer

surface has been first removed. It should then be laid upon a slide

with a drop or two of water, and covered with a piece of thin glass.

Often the cutting of the section seems to be such a shock to the leaf

that no motion is visible for awhile, but in a short time the warmth

of an ordinary sitting-room will revive it, and with a quarter-inch

object-glass the currents will be rendered beautifully distinct.

Where the “stream” is unusually obstinate the warmth may be slightly

increased, but too great heat destroys the movement altogether. In the

summer, any of the leaves show this “circulation” very well; but in the

winter, the slightly yellow ones are said to be the best.

The Vallisneria requires to be cut in sections to show this

“circulation;” but there are many plants of which it is but necessary

to take a fragment and lay it upon a slide. The _Anacharis alsinastrum_

is one of these: it grows in water, having three leaves round the

stem, then a bare portion, again another three leaves, and so on. One

of these leaves must be plucked close to the stem, and laid upon a

slide with a drop of water. Thin glass should be placed upon it, and

along the mid-rib of the leaves the “circulation” may be seen most

beautifully when a good specimen has been chosen; but it requires

rather more power than the Vallisneria. This plant is very common in

many parts of the country, a great number of our ponds and streams

being literally choked up by it. In the _Chara vulgaris_ and two or

three of the Nitellæ, &c., this phenomenon may also be seen with no

preparation except plucking a part from the stem and laying it upon a

slide as with the Anacharis. In using the Frog-bit, the outer part of

the young leaf-buds must be taken to obtain the best specimens for this

purpose; but a section of the stem will also show the “circulation,”

though not so well. The plants before mentioned are all aquatic, but

the same movement of the globules has been observed in several kinds

of land plants, as in the hairs upon the leaf-stalks of the common

groundsel; but these do not show it so well, nor are they so easily

managed as the above.

Many microscopists who are not fortunate enough to be in the

neighbourhood of these plants (indeed the Vallisneria is a foreign one)

grow them in jars, so a few remarks as to the treatment they require

will not be out of place. The Vallisneria requires a temperature not

lower than 55° or 60°, and even a higher degree than this renders its

growth quicker; and no great change must take place: the more equable

the temperature the more healthy will the plant be. A glass jar should

be taken, having an inch or two of mould at the bottom, which must be

pressed down closely, and the plant must be set in this. Water must

then be gently poured in, so as not to disturb the mould. As this plant

flourishes best when the water is frequently changed, Mr. Quekett

recommends that the jar should be occasionally placed under a tap of

water, and a very gentle stream allowed to fall into it for several

hours, by which means much of the confervoid growth will be got rid of

and the plant invigorated. The Anacharis may be rooted in the earth

like the Vallisneria, but a small detached piece may be thrown into the

jar of water and there left until wanted. For months the “circulation”

will be well shown by it, and it will probably grow and increase. It

is also very healthy in an in-door aquarium. It is recommended that

the jars in which any of the _Chara_ are grown should be moved about

as little as possible, as the long roots are very tender, and will not

bear agitation.

An object which is interesting to the microscopist, as well as the

unscientific observer, is the _growth_ of seeds, as it is often

erroneously termed. A shaving of the outside of the seed is taken and

laid upon the glass slide; a thin glass cover is then placed upon it,

and a drop of water applied to the edge of this. The water will then

gradually flow under the glass and reach the section of the seed,

when the transparent fibres will appear to spring out and “grow” for

some minutes. This, however, is produced by the unfolding of a spiral

formation in the cells, and, therefore, has really no similarity to the

true growth. The seeds of the Salvias, Collomias, Senecio, Ruellia,

&c., are well suited for the display of this curious sight.

To watch the development of the spores of ferns, and the fertilization

and products, Dr. Carpenter recommends the following mode of

proceeding:--“Let a frond of a fern, whose fructification is mature,

be laid upon a piece of fine paper, with its spore-bearing surface

downwards; in the course of a day or two this paper will be found

to be covered with a very fine brownish dust, which consists of the

discharged spores. This must be carefully collected, and should be

spread upon the surface of a smoothed fragment of porous sandstone;

the stone being placed in a saucer, the bottom of which is covered

with water, and a glass ‘tumbler’ being inverted over it, the

requisite supply of moisture is insured, and the spores will germinate

luxuriantly. Some of the prothallia soon advance beyond the rest;

and at the time when the advanced ones have long ceased to produce

antheridia, and bear abundance of archegonia, those which have remained

behind in their growth are beginning to be covered with antheridia.

If the crop be now kept with little moisture for several weeks and

then suddenly watered, a large number of antheridia and archegonia

simultaneously open, and in a few hours afterwards the surface of

the larger prothallia will be found almost covered with moving

antherozoids. Such prothallia as exhibit freshly opened archegonia are

now to be held by one lobe between the forefinger and thumb of the

left hand, so that the upper surface of the prothallium lies upon the

thumb; and the thinnest possible sections are then to be made with a

narrow-bladed knife perpendicularly to the surface of the prothallium.

Of these sections, which after much practice may be made no more than

1-15th of a line of thickness, some will probably lay open the canals

of the archegonia, and within these, when examined with a power of 200

or 300 diameters, antherozoids may be occasionally distinguished.”

Another interesting object to the young microscopist is afforded by the

spores of the equiseta (or horsetails, as they are often called). These

may be obtained by shaking the higher portion of the stems when the

spores are ripe. They will then fall like small dust, and may be placed

under the microscope. The spores are then seen to consist of a somewhat

heart-shaped mass with bands rather intricately curled around it. As

they dry these bands expand, and are seen to be four lines at right

angles, with the ends clubbed, as it may be called. If, whilst watching

them, the spores are breathed upon, these bands immediately return to

their former state, and are closely curled around the spore; but as

they gradually dry again expand. This experiment may be repeated many

times, and is a very interesting one.

The above are the principal objects which could not possibly be

included in any of the former chapters, but would have left a most

interesting branch untouched had it been neglected. There is another

subject also which should not be passed by--viz., the production of

minute pictures which serve as objects for microscopic examination.

I may here mention that as this manual is simply to enable the

young student to prepare and mount his objects, the photography of

_magnified_ objects has evidently no place here.

Few slides caused so much astonishment as these minute photographs when

first exhibited; small spots were seen to contain large pictures, and a

page of printed matter was compressed into the one-hundredth part of a

square inch. It would be impossible in this place to give the inquirer

any instruction in the manipulation of photography, so it must be

assumed that he already knows this.

We will first consider the process performed by artificial light. The

collodion employed in photographing generally shows as much structure

when magnified as is found in linen of moderate texture; but this is

not always the case, as some samples bear much enlargement without any

of this appearance. It is evident that a structure so coarse would make

it entirely unfit for these minute pictures, as all the small markings

would be destroyed, or so interfered with that no great enlargement

would be practicable. To obtain almost structureless collodion is not

an easy matter, and a clever practitioner in this branch of photography

states that he knows of no method to accomplish this with certainty,

but he himself tries different samples until he falls upon a suitable

one, which he then lays aside for this object. A beneficial effect

is often derived from keeping the collodion awhile, but this is not

always the case. The slides should be chosen of an equal thickness,

so that when focussed upon one no re-adjustment may be necessary for

the others. The glass should, of course, be free from any roughness,

scratches, or other imperfections, and of first-rate quality and colour.

The microscope must then be placed in a horizontal position, and the

eye-piece removed, the stage having a small clip upon it to keep the

prepared plate in position. The negative must then be supported at a

distance from the end of the microscope tube from which the eye-piece

was withdrawn. This distance will, of course, vary according to the

relative sizes of the negative and desired picture. With a one-inch

object-glass, which is a very convenient focus, it will have to be

changed usually betwixt one and four feet. The negative must be lighted

by an argand gas-burner or camphine-lamp, and the rays rendered as

parallel as possible by the use of a large lens placed betwixt the

light and the negative. It is not easy to arrange the apparatus so as

to get the light _uniform_; but a little practice will soon do away

with this difficulty. Ordinary ground-glass is too coarsely grained

to focus upon, as the magnifying power used to examine the minute

reflection must be considerable. One of the slides must therefore be

coated with the collodion, submitted to the silver-bath, and after

washing with water be allowed to dry. Upon this may be focussed

the reflected image, and its minuteness examined with a powerful

hand-magnifier, or another microscope placed behind in a horizontal

position. When the utmost sharpness of definition is obtained, it

is usually required to remove the plate a little distance from the

object-glass, as object-glasses for the microscope are slightly “over

corrected,” and the chemical rays which accomplish the photography

are beyond the visual ones. The exact distance required to give a

picture to bear the greatest enlargement cannot be given by rule; but

experiments must be made at first, and it will always be the same with

the object-glass which we have tested.

The plate may now be prepared as in ordinary photography, and placed

upon the stage whilst the light is shaded. When all is ready, the

shade is removed and the process allowed to go on, usually for thirty

or forty seconds; but no certain rule can be given as to the required

time, on account of various collodions, lamps, and powers being used.

It may be here mentioned, that it is well to contrive some little frame

to receive the prepared plate, as the silver bath solution is liable to

get upon the microscope stage and so, to say the least, disfigure it.

When the exposure has been continued sufficiently long, the picture

may be developed by any of the ordinary methods, but some of the best

productions have been brought out by the aid of pyrogallic and citric

acid solution, with the addition of a little alcohol. The “fixing” may

be effected by a strong solution of hypo-sulphite of soda, and the

picture must then be very well washed with pure water. When dry, the

photograph must be mounted with Canada balsam, in the same manner as

any ordinary object; but great heat must not be used, or the picture

may be injured.

When ordinary daylight is employed for this purpose, a dark slide

will be required for the prepared plate, in the same way as for

photographing landscape, &c. These dark slides are generally made by

each individual to suit his particular arrangements of negatives, &c.;

but it may be here recommended that the operator should always focus in

the same slide which he is about to use, as so small a difference in

distance lies betwixt perfection and failure.

For an ordinary student, perhaps the above method is that which is the

most readily used, and consequently the most generally available; but

almost every one has a different arrangement of microscope, &c., by

which he procures these minute pictures. Mr. Shadbolt (one of our most

successful photographers) gives the following instructions:--“Having

removed the upper stage plate of a large compound microscope, I replace

it with one of wood, supplied with guide pins of silver wire, in order

to admit of its supporting a slip of glass coated with collodion, and

excited in the nitrate of silver bath in the usual way. If the ordinary

brass stage plate were left undisturbed, it is obvious that it and the

excited slip of glass would be mutually destructive.

“The microscope is now to be placed in a horizontal position, the

objective, intended to produce the picture, made to occupy the place

usually filled by the achromatic condenser on the _sub-stage_ of the

microscope, while _another_ objective is screwed into the lower end of

the body of the instrument, which is used not only to focus with, but

also to make the requisite allowance for actinic variation.

“The negative intended to be reduced is then arranged vertically, with

its centre in the axis of the microscopic body, at a distance of from

two to four feet from the lower object-glass, and with a convenient

screen of card, wood, or thick paper, to cut off any extraneous light

that would otherwise pass beyond the limits of the picture.

“A small camphine-lamp is employed for the purpose of illuminating the

negative, having a good bull’s-eye lens as a condenser, so arranged

with its flat side next the lamp that the refracted rays shall just

fill the whole of a double convex lens of about six inches in diameter,

the latter being placed in such a position as to refract the rays of

light in a parallel direction upon the negative. By this arrangement

the _bull’s-eye lens_ of about two inches and a half in diameter

_appears_ as the source of the light instead of the small flame of the

lamp.

“By using a bat’s-wing gas-burner of a good size, a _single_ lens,

instead of the two, may be so placed as to give the necessary

uniformity of illumination.”

This arrangement requires the same care in working as that before

mentioned, the pictures being produced, developed, and fixed by the

same treatment.

As before stated, almost every manipulator makes some small changes in

the method of producing these minute pictures; but the rules given,

though far from new, are sufficient for all purposes; and I may state

with truth, that those which I procured when these wonders were quite

new, are fully equal in every respect to the best usually met with at

the present time.

With these instructions I shall close my Handbook, as I believe that

every branch of Preparation and Mounting of Microscopic Objects has

been treated of. Not that the beginner can expect that he has nothing

to do except read this to be able to mount everything; but there are

difficulties from which he may be freed by instruction, when otherwise

he would have been compelled to learn by failure alone. I may, here,

however, repeat certain advice before given,--that, when practicable,

it is a good thing to mount each object by two or more different

methods, as very frequently one feature is best shown dry, another in

liquid, and a third in balsam. Secondly, let the _mounting_ be studied

thoroughly, as no part of the microscopic science is more worthy of

thought than this. And lastly, let no failures prevent you following

up what will assuredly one day become a source of great pleasure, and

render your daily “constitutional walk,” which is often dull in the

extreme, very delightful, as it will afford you some new wonder in

every hedge-row.

INDEX.

Air-bubbles, 56.

Air-pump, 13;

use of, 57.

Alcock, Dr., on tongues of Mollusca, 118.

Algæ, mounting of, 91.

Anacharis alsinastrum, rotation in, 143;

to cultivate, 144.

Animal tissues, dissection of, 114.

Antennæ of insects, 71;

in preservative liquid, 94.

Apparatus required in mounting objects, i.

Arteries, how to distinguish, in injection, 131.

Asparagus, spiral vessels of, 114.

Asphaltum, 15;

and india-rubber, as cement and varnish, 18.

Bat, circulation of blood in wings of, 142.

Beale’s, Dr., Prussian blue for injecting, 134;

carmine, 135.

Bell-glasses, use of, 12.

Berg-mehl, 40.

Bermuda earth, 40.

Bichromate of lead, injection with, 132.

Bird, Dr. Golding, on preparation of Zoophytes, 65.

Black-japan, as a cement, 16.

Black varnish, 18.

Blood, as a microscopic object, 50;

circulation of, 140–142.

Bone, sections of, 102;

fossil, 103.

Brass plate for heating glasses, &c., 13.

Cactaceæ, raphides of, 52.

Camel-hair pencils, use of, 10.

Canada balsam, 14;

with chloroform, 14, 60, 132;

with turpentine, 14;

air bubbles in, 56;

Dr. Carpenter’s syringe for, 59;

to fill cells with, 60;

mounting of objects in, 56.

Carbolic acid, 86.

Carmine injecting fluid, Dr. Beale’s, 135:

Dale & Davies’, 138.

Cartilage, dissection of, 114.

Castor oil, as a preservative, 86.

Cells for dry objects, 6;

with rings of cardboard, 7;

with gutta-percha, 7;

with leather, 7;

with ivory, 8;

with thin glass, 7;

with varnish, 23, 26;

for balsam, 60;

for preservative liquids, 87–89;

Shadbolt’s turntable useful in making, 9.

Cements, 13–17.

Chalk, foraminifera from, 63.

Chara vulgaris, rotation in, 144.

Chloride of zinc, solution of, 85;

of calcium, solution of, 92.

Chloroform, use of (_see_ Canada balsam).

Circulation of blood, 140–142;

of sap (_see_ Rotation).

Cleanliness in microscopic work, 1.

Coal, sections of, 99.

Collection of diatoms, 29.

Colours for injection, 125–127.

Condenser, cheap, to make, 20.

Corals, sections of, 99.

Corallines, to mount, 48.

Cover of objects, to remove, 60.

Crystals, mounting of, dry, 50;

to vary form of, and mount in balsam, 75;

sections of, 109.

Cuticle of equisetum, &c., 80.

Dale & Davies’, carmine injection, 138.

Deane’s gelatine, 84.

Decalcifying process for shells, 98.

Desmidiaceæ, in preservative liquid, 92.

Diachæa elegans, 53.

Diamond beetle, 54.

Diatomaceæ, nature of, 28;

collection of, 29;

in stomachs of fish, 30;

to prepare and mount dry, 30–40;

Mr. Rylands on, 33;

in guano, 39;

fossilized, 40;

mounting in balsam, 61;

mounting in preservative liquids, 92, 93.

Discs used in mounting, 22;

supporter, Smith & Beck’s, 23.

Dissection, 111;

microscope for, 111;

instruments for, 112.

Distilled water, as preservative liquid, 83.

Doyer’s, M., method of injection, 132.

Dry objects, to mount, 22.

Echinodermata spines, sections of, 99.

Eel, scales of, 78.

Eggs of insects, 94.

Elastic objects, to keep flat upon the slide, 11.

Electrical cement, 16.

Equisetaceæ, 80;

spores of, 146.

Erector, not indispensable, 19.

Eyes of insects, 70, 54;

of animals injected, 136.

Feathers, 73.

Feet of insects, 54, 71.

Ferns, 51;

development of spores of, 145.

Fish, fins and tails of, 50;

scales of, 53, 78;

injection of, 137;

circulation of blood in, 142.

Flint, sections of, 101.

Fluid, mounting objects in, 83.

Flustra avicularis, 79.

Fly (_see_ Insects).

Foot of sheep injected, 136.

Foraminifera, 41;

separation of, 41;

to clean, 42;

to clean from tallow soundings, 42;

to mount, 45, 63;

from chalk, 63.

Forceps, ordinary and wooden, 11;

bull-nosed, for injection, 123.

Fossil infusoria, 40.

Frog, injected, 136;

to show circulation of blood, 140.

Frog-bit, rotation in, 144.

Fruit-stones, sections of, 104.

Fungi, 53, 94.

Gastric teeth of insects, 120;

of molluscs, 117.

Gelatine for injection, 124;

preservative liquid, 84.

Glass slides, 1;

thin, 3;

thin, to cut, 3;

thin, to measure, 4;

thin, to clean, 5;

cells, to make, 88;

rings for cells, 88;

tubes, 10.

Glycerine, 84, 94;

jelly, 84, 91.

Goadby’s fluid, 85.

Gold-size, 15.

Grasses, 80.

Grasshopper, gizzard of, 120.

Guano, containing Diatomaceæ, 39.

Gum-water, and modifications of, 17.

Gutta-percha cells, 7;

for liquids, 87.

Hairs, vegetable, 46;

to mount dry, 54;

to mount as polarizing objects, 79;

sections of, 105.

Hepworth, Mr., on mounting insects, 68.

Horn, sections of, 104.

Hot-water bath, use of, 58.

Infusoria, in preservative liquid, 92;

fossil, 40.

Injections, vi., 122;

apparatus for, 122;

colours for, 125;

directions for, 127;

with various colours, 131;

mounting of, 131;

transparent, 133, 138.

Insects, scales of, 48;

to mount, 49;

legs and feet of, 54, 72;

eyes of, 54, 70;

Mr. Hepworth on mounting, 68;

antennæ of, 71;

mouth of, 72;

tracheæ and spiracles of, 72, 115;

parasitic, 73;

in preservative liquid, 94;

eggs of, 94;

gizzard of, 120;

circulation of blood in, 142.

Intestines, injected, 136.

Knives for dissecting, 112;

Valentin’s, 108.

Labelling of objects, 19.

Lamps, for mounting, 12.

Larvæ, skins of, 50.

Leaves, sections of, 107;

scales of, 46, 81.

Liquid-glue, 16.

Lungs of animals injected, 137.

Mallow, pollen of, 47.

Marine glue, 15, 88.

Microscope for dissection, 111.

Miscellaneous, vii., 140.

Mites, 73.

Molluscs, tongues of, 116.

Mosses, 51;

in preservative fluids, 91.

Mould (_see_ Fungi).

Mounting objects, apparatus for, i.;

dry, ii., 22;

in Canada balsam, iii., 56;

in cells, iv., 83.

Mouse, ear of, 114;

circulation of blood in, 142.

Mouth of insects, 72.

Muscle, dissection of, 114.

Needles, how to mount, 10;

for dissection, 112;

curved, for injection, 123.

Nervous tissue, dissection of, 115.

Nettle leaf, 52.

Newts, injected, 136.

Onion, raphides of, 52.

Orbitolite, section of, 97.

Oxalurate of ammonia, crystals of, 75.

Palates of Molluscs (_see_ tongues).

Papers, ornamental, to cover slides, 8, 27.

Photographs, microscopic, to produce, 147;

Mr. Shadbolt on, 149.

Pipes for injecting syringe, 122.

Podura, scales of, 49.

Polariscope, objects for, 74–82, 104.

Pollen, 47, 74.

Polycystina, preparation and mounting of, 63.

Preservative liquids, iv., 83;

cells suited for, 87–89.

Prussian blue for injection, 134.

Raphides, vegetable, 52, 80.

Rhinoceros, horn of, 104.

Rhubarb, spiral vessels of, 113.

Rings and cross of crystals, 109.

Rotation of fluid in cells of plants, 143–145.

Rush, section of, 108.

Rylands, Mr. T. G., on Diatomaceæ, 33, 93.

Salicine, crystals of, 76.

Saw of watch-spring, 97.

Scales of fishes, 53, 78;

of leaves, 46, 81;

of insects, 48.

Scissors, 10;

for dissection, 112.

Sea-mats, 53.

Sea-soundings, to cleanse, 42.

Sealing-wax varnish, 17.

Sections, 96;

of shells, 97;

of orbitolite, 97;

of spines of Echinodermata, 99;

of corals, 99;

of coal, 99;

of flint, 101;

of teeth, 101;

of bone, 102;

of fruit-stones, 104;

of horn, 104;

of whalebone, 105;

of hairs, 105;

of wood, 106;

of leaves, 107;

of sponges, 108;

of skin, 109;

of crystals, 109;

of seeds, 111.

Seeds, 47, 74;

sections of, 111;

growth of, 145.

Shadbolt’s turntable, 9.

Shells, sections of, 97;

decalcifying, 98;

laminæ of, 98.

Siliceous cuticles, 80.

Size for injection, 124.

Skins of larvæ, 50;

sections of, 109;

sole, 54.

Slides, glass, for mounting objects, 1;

glass, to clean, 2;

wood, &c., 6;

most useful, 8;

to cover and varnish, 27.

Spicula, from sponges, &c., 67.

Spines of Echinus, 99.

Spiracles of insects, 72, 116.

Spiral vessels of vegetables, 113.

Split bristles, use of, 10.

Sponges, sections of, 108.

Spores of ferns, development of, 145;

equisetum, 146.

Starch, preparation and mounting of, 79.

Sulphate of copper and magnesia, crystals of, 76.

Syringe for Canada balsam, 59;

for dissection, 113;

for injection, 122.

Tadpole, to show circulation of blood of, 141.

Teeth, sections of, 101.

Thin glass, to cut, 3;

to measure thickness of, 4;

to clean, 5.

Thwaites’ preservative liquid, 85.

Ticks, 73.

Tissues, animal and vegetable (_see_ Dissection).

Tongues or palates of Molluscs, 116.

Tracheæ of insects, 72, 115.

Transfer of objects, 10.

Trough for dissection, 113.

Tubes, glass, 10.

Turnbull’s, Dr., Prussian blue for injection, 134.

Turpentine, use of, 57.

Universal stand, to make, 20.

Valentin’s knife, 108.

Vallisneria spiralis, rotation in, 143;

to cultivate, 144.

Varnishes, 17.

Vegetable objects, to mount dry, 46;

to mount in jelly, 91;

dissection of, 113.

Watch-glasses, 12.

Whalebone, sections of, 105.

Wood, sections of, 106.

Zoophytes, to mount dry, 53;

Dr. Golding Bird on mounting, in balsam, 65;

as polarizing objects, 79.

COX AND WYMAN, PRINTERS, GREAT QUEEN STREET, LONDON.

Transcriber’s Notes

Punctuation and spelling were made consistent when a predominant

preference was found in the original book; otherwise they were not

changed.

Inconsistent hyphenation was not changed.

Simple typographical errors were corrected; unbalanced quotation

marks were remedied when the change was obvious, and otherwise left

unbalanced.

The Table of Contents was added by the Transcriber.

The index was not checked for proper alphabetization or correct page

references. Roman numeral references are to chapters, not to pages.

Page 106: =T= indicates a boldface, sans-serif “T”.