414 SAMUEL C. PALMER 
successful double staining, to which I shall refer later, whereby 
small fragments of rods could be distinguished from bits of cones. 
The large size of all the retinal cells in Necturus has been a great 
aid in the identification, and in the accuracy of the counts, of 
the separate elements of the retina. The most successful re- 
sults were obtained by fixation in Kleinenberg’s picro-sulphuric 
mixture. My method was as follows: Live animals were placed 
in a bowl of tapwater in which a few small crystals of chloretone 
had been dissolved as a means of preventing the discharge 
of slime (Cole, ’02). Chloroform was then added gradually 
until the animals were thoroughly anesthetized. The eyes were 
quickly removed and placed in picro-sulphuric acid, care being 
taken to free them from as much superfluous tissue as could be 
done quickly. To keep the orientation, a piece of skin was left 
attached to the dorsal side of the eyeball, differences in shape 
of the pieces serving to distinguish right and left eyes. I found 
the pieces of skin useful also in orientating the eyes in paraffin. 
The best results were obtained by immersing the unopened eye- 
balls in the picro-sulphuric mixture for four or five hours. ‘They 
were then rinsed in distilled water a few minutes and dehydrated 
by passing them gradually through 35 per cent, 50 per cent, 90 
per cent, and 100 per cent alcohol over a period of two days. 
When the eye was sufficiently hardened (90 per cent alcohol), 
the front face was cut away with a sharp razor, and the lens 
removed. Early in my work I found that the heat of a paraffin 
bath, extending over a time sufficient to insure saturation with 
paraffin, caused considerable shrinkage of the sclera and wrink- 
ling of the retina. I, therefore, followed Biitschli’s chloroform 
method of de-aleoholization. Transfer from the chloroform-par- 
affin mixture of this method to hard paraffin was completed by 
the evaporation of the chloroform over a water bath at about 
60°C. and a five minute immersion in hard paraffin melting at 
about 56°C. 
Sections 8» thick were made in one set of eyes parallel to the 
antero-posterior! plane of the eye and passing through the optic 
1 The terms ‘anterior,’ ‘posterior,’ ‘dorsal’ and ‘ventral’ as applied to the eye- 
ball in this account, are used in the sense of comparative anatomy; i.e.. ‘anterior’ 
