360 
PACIFIC SCIENCE, Vol. V, October, 1951 
Fig. 1. Dissection of a mature male Panulirus penicillatus (Oliver) drawn to show the structure and position 
of the reproductive system in relation to other structures, a. Intestine; b, testis just anterior to transverse bridge; 
c, pyloric region of stomach; d, enlarged portion of vas deferens; e, hepatopancreas. (0.5 X) 
METHODS AND TECHNIQUES 
Specimens of P. penicillatus taken in the 
vicinity of Kaneohe Bay, Oahu, between 
July, 1947 , and January, 1948, were used in 
this study. Males whose carapace length ex- 
ceeded 10 centimeters were usually found to 
possess well-developed gonads. These were 
removed from freshly killed specimens, rolled 
lightly on blotting paper, weighed, and 
placed immediately in fixative. A label indi- 
cating the catch and specimen number was 
inserted in each vial so that histological data 
could be correlated with size, date, and other 
pertinent information. 
For the purpose of routine histological 
examination the reproductive system (Figs. 
1, 2) was divided into: anterior testis (Fig. 
2/); mid-testis {e) \ posterior testis (c); proxi- 
mal vas deferens {d)\ and the massive distal 
vas deferens {h). Small portions of these 
regions were placed in Bouin’s fixative, which 
gave excellent preparations for all tissue ex- 
cept the vas deferens which became extremely 
brittle. Tissues were cleared with toluene, 
embedded in Tissuemat (54-56° C.) and, for 
routine investigation, sectioned at 10 mi- 
crons. Some testis preparations were placed 
in Bouin’s fixative and some in Fleming’s, 
and were cut at 6 and 4 microns. Routine 
sections were stained with standard alum- 
haematoxylin and counterstained with eosin 
(0.5 per cent solution in 90 per cent alcohol 
to which 04. cc. of 0.1 N HCl was added). 
Heidenhain’s iron-haematoxylin without a 
counterstain was used with good results in 
some sections of the testis. Mallory’s triple 
connective-tissue stain was employed to aid 
in tissue differentiation. Because of the ex- j! 
treme hardness of the enlarged portions of 
the vas deferens, it was often necessary to ; 
coat the surface of the paraffin block with j 
thin celloidin between successive cuts. For || 
examination of the rayed spermatozoa, fresh ; 
sections were taken from the enlarged vas j 
deferens, the matrix was separated from the 
spermatophore, and the spermatophore was 
opened in sea water. j 
Figures 1 and 9 were drawn by Florence i 
Lambeth from dissections made by the I 
author. Figures 2, 4, 5, 6, 7, and 8 were drawn 
by Inger Achton from slides and dissections 
made by the author. Figure 3 was drawn by 
Evan L. Gillespie. 
The writer acknowledges with thanks the 
help of Dr. Robert W. Hiatt, who suggested 
the problem and offered his assistance in many 
ways throughout the course of the study. 
b c 
