Fluorescent enzyme-based biosensor for sensitive analysis of DNA damage in cryopreserved sperm

Area of Science:

• Reproductive Biology
• Biotechnology
• Sperm Cryopreservation

Background:

• Freeze-thaw cycles cause irreversible sperm DNA damage, impacting fertility.
• Accurate assessment of sperm DNA integrity is vital for utilizing cryopreserved sperm effectively.

Purpose of the Study:

• To develop and validate a novel fluorescent biosensor for detecting DNA breakpoints in human sperm after cryopreservation.
• To compare the biosensor's efficacy against the conventional sperm chromatin structure analysis (SCSA) for DNA fragmentation index (DFI) measurement.

Main Methods:

• A novel fluorescent biosensor utilizing terminal deoxynucleotidyl transferase (TdT) and Endonuclease IV was employed.
• Sperm DNA integrity was assessed in fresh, cryopreserved, and antioxidant-treated groups.
• Biosensor results were compared with DFI measurements from SCSA.

Main Results:

• Cryopreserved sperm showed significantly higher DFI than fresh sperm.
• The novel biosensor detected a significant reduction in DNA breakpoints in the antioxidant group compared to the cryopreserved group.
• The biosensor demonstrated superior accuracy and effectiveness over SCSA in assessing sperm DNA integrity post-cryopreservation.

Conclusions:

• The novel fluorescent biosensor is a highly accurate and effective tool for evaluating sperm DNA integrity after cryopreservation.
• This biosensor offers significant potential for enhancing the clinical and scientific application of cryopreserved sperm in reproductive medicine.