A conditioned medium (CM) was prepared by culturing human sperm at 3.0 x 10(6) sperm/ml for 12 h. The fusion index increased significantly when the sperm penetration assay (SPA) was performed with CM (0.6 without CM to 2.0 with CM). The conditioned medium was also effective on the sperm from 11 patients who showed a low or zero value in SPA and who's in vitro fertilization treatment failed. When CM from the sperm of healthy donors was added to these patients' sperm, the fusing ability to zona free hamster eggs was significantly restored.