The purpose of this study was to compare the different sperm manipulation methods on the recovery rate of motile sperm that was collected to use in the intrauterine insemination. Three hundred and thirty-five IUI (intrauterine insemination) cycle from a total of 963 treatment cycles was performed for non-peritoneal factor infertility couples. The sperm separation methods were swim-up, swim-down and Percoll gradient (since Mar. 1992). We compared the different sperm preparation methods in normal and abnormal semen analysis group. Our results show that in the abnormal semen samples (oligospermic approximately, asthenospermic approximately), the Percoll gradient method is superior to the swim-up and swim-down methods on the recovery of a higher number of total, motile and active motile spermatozoa (p < 0.05), even though the sperm quality before preparation was rather poorer than the samples prepared by the other two methods. The pregnancy rate in both groups are not statistically significant. We conclude that the Percoll gradient is an improved method for semen manipulation as it allows greater recovery of sperm with higher motility in intrauterine insemination.