Background: Sperm cryopreservation is currently the only clinically available technique for preservation of male gametes for subsequent use in assisted reproduction technologies. Objective: The objective of this study was to determine the impact of semen cryopreservation on human sperm DNA. Patients and methods: The study sample consists of 20 male patients seeking seminal fluid analysis at the andrology laboratory of a specialized IVF center (ADAM International Hospital for Fertility and Sterility, Giza, Egypt), and accepting to sign a consent of approval to participate in the study, with the diagnoses of normozoospermia, oligozoospermia (either isolated or combined with asthenozoospermia or teratozoospermia). Results: There is no statistical difference between Pre-freezing and vitrification regarding DNA fragmentation, P>0.05. No statistical difference between Pre-freezing and slow frozen regarding DNA fragmentation, P>0.05. Finally, there is no significant statistical difference between vitrification and slow frozen regarding DNA fragmentation, P>0.05. Conclusion: The sperm DNA fragmentation index was not affected during cryopreservation under the various methods of storage tested. Clinicians and investigators should take this information into consideration when using cryopreserved sperm for assisted reproduction.