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.