Evaluation of Sperm DNA Fragmentation Using Halosperm Technique after the Freezing–Thawing Process in Men: A Study on the Validation of the SCD Protocol

Chaymae  Rochdi; Larbi  Allai; Ibtissam  Bellajdel; Hafsa  Taheri; Hanane  Saadi; Ahmed  Mimouni; Mohammed  Choukri

doi:10.18502/jri.v25i1.15194

Chaymae Rochdi Maternal-Child and Mental Health Research Laboratory, Faculty of Medicine and Pharmacy, Mohammed First University, Oujda, Morocco
Larbi Allai Laboratory of Sustainable Agriculture Management, Higher School of Technology Sidi Bennour, Chouaib Doukkali Uni-versity, El Jadida, Morocco
Ibtissam Bellajdel Medically Assisted Procreation Unit, Central Laboratory, Mohammed VI University Hospital Center, Oujda, Morocco
Hafsa Taheri Maternal-Child and Mental Health Research Laboratory, Faculty of Medicine and Pharmacy, Mohammed First University, Oujda, Morocco
Hanane Saadi Maternal-Child and Mental Health Research Laboratory, Faculty of Medicine and Pharmacy, Mohammed First University, Oujda, Morocco
Ahmed Mimouni Maternal-Child and Mental Health Research Laboratory, Faculty of Medicine and Pharmacy, Mohammed First University, Oujda, Morocco
Mohammed Choukri Maternal-Child and Mental Health Research Laboratory, Faculty of Medicine and Pharmacy, Mohammed First University, Oujda, Morocco

DOI: https://doi.org/10.18502/jri.v25i1.15194

Keywords: Cryopreservation, DNA fragmentation, Freezing, Halosperm test, Spermatozoa, Thawing.

Abstract

Background: DNA fragmentation index (DFI) enhances routine semen analysis by providing valuable insights into male reproductive potential. Utilizing Halosperm test, a sperm chromatin dispersion (SCD) assay based on induced condensation. The purpose of this study was to assess sperm DNA damage both before and after freezing. By following the specified kit instructions, an attempt was made to validate the SCD test protocol, with a particular emphasis on the implications of sperm freezing on its DNA integrity.

Methods: In total, 380 fresh human semen samples from normozoospermic patients were frozen at -20°C for 10 days, using SCD cryopreservation reagent. Routine semen analysis and DNA fragmentation index (DFI) were determined for each sample before freezing and after thawing. Semen morphology and sperm DFI were compared before and after freezing/thawing process.

Results: There was a significant decrease in sperm normal morphology after thawing (9.31±2.42% vs. 7.1±1.53%, p<0.05, respectively). The sperm head, midpiece, and tail defect rate increased after freezing at -20°C. Moreover, DFI was significantly higher after thawing compared to before freezing (20.71±1.61% before freezing vs. 29.1±0.21% after thawing with p<0.001).

Conclusion: Cryoconservation of semen samples at -20°C for 10 days using SCD cryopreservation reagent seems to damage sperm morphology, resulting in a reduction in sperm DNA integrity. The measurement of DFI on a fresh sample remains the most reliable technique for obtaining accurate results.