With 1% lecithin and 1 ·0 m-glycerol, the activity of deep-frozen bull spermatozoa after thawing was better if they were frozen in an isotonic diluent than if frozen in a hypertonic ( 1 ·2 x ) solution (the solution referred to as isotonic was : 50 mm-sodium citrate, 62 mm-fructose and 20 mm-sodium phosphate buffer). There was no significant difference between the mean effects of addition of the glycerol either immediately the diluted semen had been cooled to 5° C (equilibration) or just prior to freezing after a period of storage at 5° C equivalent to the equilibration period (ageing). A period of 4 hr of storage at 5° C prior to freezing gave the best mean motility scores and percentage motile, but the percentage of spermatozoa unstained by congo-red-nigrosin solutions continued to rise over a period of 18 hr storage.
Yolk-containing diluents, with the tonicity range of 0·90, 1·05 and 1·20 x, were tested and hypotonic diluents were superior to hypertonic. Equilibration for up to 18 hr had no significant effect on the scores of activity of spermatozoa after thawing, but, again, there was a highly significant rise in the percentage of unstained spermatozoa.