Variation in ejaculatory abstinence time and its influence on semen quality and clinical reproductive outcomes is a growing concern among clinicians and researchers. The WHO (World Health Organization) recommends 2–7 days of abstinence time prior to semen collection for diagnostic purposes; however, the evidence that such an abstinence period leads to better pregnancy outcomes remains unclear. The aim of this systematic review is to evaluate short and long ejaculatory abstinence time in association with pregnancy rate, live birth rate and DNA fragmentation, in order to make a recommendation on an ideal timeframe for ejaculatory abstinence. This review is conducted according to the PRISMA guidelines and registered in PROSPERO (CRD42022379039). The electronic databases PubMed, Embase and Cochrane were searched for eligible studies. The Scottish Intercollegiate Guidelines Network was used for the assessment of the risk of bias across the included studies. Twenty-four studies were included in this systematic review. The included studies confirm that a shorter abstinence time is associated with improved pregnancy rates and live birth rates following assisted reproductive technology compared with longer ejaculatory abstinence times at different cut-off points. Similarly, a lower DNA fragmentation index was reported in semen analyses collected from short abstinence times compared with long abstinence times. However, due to the heterogeneity of the included studies, it is not possible to extract an ideal time of ejaculatory abstinence, but all outcomes improved with shorter ejaculatory abstinence times. This systematic review confirms that short ejaculatory abstinence times, less than those recommended by the WHO for diagnostic purposes, are associated with higher pregnancy and live birth rates and improved DNA fragmentation, when compared to long ejaculatory abstinence times.