Background: The number of sperm washes to maximize outcomes for intra-uterine insemination has not been well investigated. Therefore, we undertook to compare the pregnancy and live birth rates of triple sperm washing and double sperm washing for density gradient preparation for intrauterine insemination (IUI) cycles. Methods: A retrospective cohort study including 279 couples (136 couples with triple sperm washing and 143 couples with double sperm washing) with a diagnosis of unexplained infertility and mild male subfertility who had IUI cycles between April 2015 and April 2017 were evaluated. After overnight incubation of the sperm, subjects underwent either traditional double sperm washing or Triple sperm washing which consists of use of a third gradient and spinning procedure to the conventional double gradient sperm washing in order to obtain a higher quantity of motile sperm. Results: Total sperm count after triple washing was higher than double sperm washing (98.25 \pm 62.06 vs. 81.08 \pm 31.57; p = 0.003). Positive \beta-hCG test and live birth per cycle were higher in triple sperm washing (25.8% vs. 13.3%, p = 0.009; 18.4% vs. 9.8%, p = 0.039; respectively) than in the double sperm washing group. Conclusions: The use of motile sperms obtained from triple sperm washing may increase the rates of pregnancy and live birth in IUI cycles of women with unexplained and mild male factor infertility. A prospective randomized study should be undertaken to confirm the results.