They may be left in this from three days to a week with little fear of overstaining. Portions may then be teased with needles, and mounted in glycerine, or the stained tissue may be pressed out into a sufficiently thin layer by squeezing it forcibly between two glass slides.
Motor and sensory nerve endings.—These are best stained by the chloride of gold method (p. [82]).
Specimens must be taken from the body immediately after death. The method is therefore useless for the post-mortem room, but may be used for tissues removed by operation. Small pieces of tissue must be employed and must be stained in bulk, sections being made subsequently.
For motor nerve endings the muscle of a frog or human muscle from a limb just amputated may be taken. Specimens should be prepared after staining by teasing in preference to making sections. Mount in Farrant.
Sensory nerve endings may be conveniently studied in the cornea of a recently killed frog or rabbit, or in a freshly extirpated human eye. Tactile end organs may be studied in the lip or finger tips, taste buds in the papilla foliata of the rabbit’s tongue, and Pacini’s corpuscles are well seen in the mesentery of a thin cat.
3. Staining nerve cells.
Bevan Lewis’s aniline blue-black method:—This method is the best for demonstrating the wealth of nerve cells in the fresh cerebral cortex. The solution of aniline blue-black should be of the strength of 1 in 400, about a grain to the ounce. A piece of the cerebral cortex with pia mater attached, should be removed as soon as possible after death by parallel cuts about 1/8 inch apart, and perpendicular to the surface of the convolution, placed on the plate of the freezing microtome and just frozen—not too hard or the tissue will be brittle and will also injure the edge of the razor. As soon as a good section is obtained the razor should be plunged into a large bowl of cold water to detach the section, which is at once floated on a glass slide, and osmic acid solution, 1/4 per cent. allowed to flow over it from a pipette. This will fix the tissue elements in about two minutes. The section is again floated off into the bowl of water and thoroughly washed to free it from the osmic acid. It is then stained either on the slide, or in a watch-glass, with the aniline blue-black solution for an hour in the cold, or half-an-hour if the solution is slightly warmed. The dye is thoroughly washed away with distilled water, excess of moisture wiped off the slide with blotting paper, and the section allowed to dry under a glass bell jar. It is not practicable to dehydrate by means of alcohol as it would cause sudden shrinking of the tissues. When the section is dry a drop of Canada balsam is applied and it is covered with alcohol in the usual way. The nerve cells and their processes are stained a deep slate colour, as are the nuclei of the connective tissue cells, while the ground work of the neuroglia is faintly stained and of a neutral grey tint. This method gives beautiful results both with normal and morbid specimens.
Various other aniline dyes, indulin, methylene blue, gentian violet, have been employed in the same way, but none of them give such good or uniform results as aniline blue-black.
Hardened specimens may also be stained with aniline blue-black, but the results are not to be compared with those obtained by the fresh method. The stain is usually diffuse, but this can be improved by placing the sections for 1/2 –2 minutes in a 2 per cent. solution of chloral hydrate in distilled water after staining.