Prevention of fertilization by exposure of hamster eggs to soluble acrosin

Incubation of hamster eggs with trypsin or chymotrypsin, but not with a series of glycosidases or lipases, prevents capacitated spermatozoa from binding to the zona pellucida so that fertilization is prevented (Hartmann & Gwatkin, 1971; Gwatkin, Williams & Andersen, 1973). Inactivation of the binding sites for spermatozoa on the zona pellucida by a trypsin-like protease released by discharge of cortical granules also appears to be responsible for the zona reaction which prevents polyspermy (Gwatkin, Williams, Hartmann & Kniazuk, 1973). Acrosin is also a trysin-like protease and it can be extracted from sperm acrosomes. We therefore investigated whether the fertilization of hamster eggs is blocked by incubation with soluble acrosin. Eggs were collected from hamsters induced to superovulate and the cumuli oophori were re¬ moved with hyaluronidase as described previously (Gwatkin, Andersen & Hutchison, 1972). Groups of such cumulus-free eggs were incubated under oil in 20-µ1 drops of PBS (pH 7-3; Dulbecco & Vogt, 1954) containing various concentrations of trypsin or acrosin for 2 h at 37-5cC on a rocker (5-6 oscillations/min). The eggs were then washed in medium 199M2 (Gwatkin & Andersen, 1973) and added to 40-µ1 drops of a suspension of epididymal spermatozoa previously capacitated by incubation for 5 h in medium 199M2 containing cumulus cells (Gwatkin et al, 1972). After incuba