MOTILITY AND OTHER CHARACTERISTICS OF HUMAN SPERM CAN BE MEASURED BY COMPUTER-ASSISTED SPERM ANALYSIS OF SAMPLES STAINED WITH HOECHST-33342

Objective: To develop methods for using a DNA-specific dye to discrimi nate between motile and nonmotile sperm and static particulate matter in fresh and diluted semen, using computer-assisted sperm analysis (CA SA) with the Hamilton Thorne NOS, TOX version (Hamilton-Thorne Researc h, Beverly, MA). Design: Donor semen was divided for treatment as fixe d stained sperm (Hoechst 33342 stain; Sigma Chemical Company, St. Loui s, MO), fresh motile and nonmotile stained sperm, and unstained contro l sperm. Setting: Normal human volunteers in art academic research and medical environment. Patients: Selected healthy student volunteers. I nterventions: Delivered semen to the laboratory within 1 hour of colle ction. Main Outcome Measure: Semen quality measured by CASA. Results: Fixed or fresh human sperm stained with Hoechst 33342 dye should be di luted to less than or equal to 50 x 10(6) sperm/mL to count sperm accu rately. Motile and nonmotile sperm mere stained suitably with 5 to 10 mu g/mL of dye when diluted with a simple diluent, but the dye concent ration should be increased to 40 mu g/mL when egg yolk is in the dilue nt. Conclusions: The DNA-specific dye, Hoechst 33342, can bt, used to discriminate between motile and nonmotile sperm and other particulate matter when evaluated by CASA with instrumentation equipped with suita ble optics.