Human sperm chromosomes in zona-free hamster eggs: pretreatment of spermatozoa with calcium ionophore facilitates later examination of chromosomes

Freshly ejaculated human spermatozoa exposed to 10 μM calcium ionophore A23187 for 30 minutes quickly bound to and penetrated into zona-free hamster eggs. When these eggs were cultured for 13 hours in the presence of low concentrations of vinblastin and podophyllotoxin, then fixed and spread with the use of a cold fixation technique, haploid sets of sperm chromosomes were demonstrated. The number of spermatozoa that were attached to the egg surface at insemination influenced the success rate of chromosome demonstration. The best results (an average of about 0.7 chromosome set per egg) were obtained when insemination was controlled so that an average of seven spermatozoa attached to each egg at the end of a 15–30-minute insemination period. This improved method for demonstrating human sperm chromosomes would be useful in the analysis of paternal genetic contributions in reproduction as well as in assessing the effects of environmental genotoxic agents or conditions on spermatozoa.