416 SAMUEL C. PALMER 
made with the use of other reagents. Both the vapor of 2 per cent 
osmic acid and vom Rath’s picro-osmo-platinic-chloride-acetic 
mixture gave excellent preservation of the retinal elements, 
though the outer segments of the rods and cones were over-black- 
ened. Good preservation and successful double-staining were 
obtained by fixation in either Perenyi’s fluid, Fol’s mixture, or 
7 per cent nitric acid, followed as before with haematoxylin and 
eosin stains. 
I have already called attention to the non-medullated char- 
acter of the optic nerve fibers in Necturus, hence the usual fixa- 
tion fluids in which osmic acid is used to stain the medullary 
sheaths are ineffective for the optic nerve fibers, in this animal. 
Vom Rath’s fluid gave excellent preservation of the supporting 
and vascular tissues, but was unsatisfactory for the nerve fibers. 
Ranson’s (’09) modification of Ramén y Cajal’s silver-nitrate 
method for non-medullated nerve fibers was tried without suc- 
cess. With another modification of this method, that of Mullenix 
(09), I succeeded in staining the fibers, but the definition was 
poor. The only method tried which brought out the fibers dis- 
tinctly was a modified form of Bielschowsky’s (’03) method. In 
order to secure the necessarily rapid fixation of the proximal 
portion of the optic nerve, I cut away the tissues surrounding 
the skull, which was then split at both anterior and posterior 
ends. This permitted the formalin to enter the brain cavity 
quickly. The non-medullated fibers when impregnated by this 
method appeared in longitudinal section as sharply defined 
somewhat undulatory, brownish-black to black lines. In cross- 
section (figs. 10, 11, 12) they were irregular black spots or streaks 
in a yellowish-brown matrix. Dehydration and de-alcoholiza- 
tion were carried out as with the retina. Cross-sections of the 
optic nerve 5y thick were made close to the chiasma and as near 
as possible to the eyeball. 
The results obtained with Bielschowsky’s fluid have justified 
its use in this case. There is no doubt in my mind that the irreg- 
ular black spots and streaks referred to above are nerve fibers. 
I was fortunate in having a longitudinal section of an optic nerve 
turn up slightly so that the fibers could be seen to end as small 
