Worm Breeder's Gazette 3(2): 14b
These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.
In order to understand the mechanism of preferential fertilization by male sperm (Gazette 2, 2), I sought a vital dye to label male sperm to distinguish them from hermaphrodite sperm after mating. All the known vital dyes and fluorescent compounds available at Carnegie were fed to worms. Most labeled only the gut, but one, the lipid stain Nile Blue A, was taken up into all the tissues. The dye fluoresces in the red with green illumination and individual sperm can be readily seen. Nile Blue A labelled males are as fertile as unlabeled males and the sperm are utilized preferentially. Observation of matings by dye labelled males revealed that the sperm migrate up the uterus among the eggs and pass through the spermathecal valve. In the spermatheca they appear to accumulate preferentially against the spermathecal wall, displacing the hermaphrodite sperm to the center of the spermathecal cavity. This displacement may be the mechanism of preferential fertilization. The dye transfers rapidly from the sperm to the walls of the spermatheca, so it is not possible to follow the sperm all the way to fertilization. This presumably reflects its lipid solubility. Males are labelled by placing larvae on seeded plates to which 0.4 ml of 0.65% Nile Blue A was added and allowed to soak in. The dye is dissolved initially at 5% in 95% ETOH and diluted With M-9 buffer prior to use.