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Volume 12, Issue 5 (September 2010) 12, 753–759; 10.1038/aja.2010.46

Preparation and incubation conditions affect the DNA integrity of ejaculated human spermatozoa

Rieko Matsuura, Takumi Takeuchi, Atsumi Yoshida

The Reproduction Center, Kiba Park Clinic, Tokyo 135-0042, Japan

Correspondence: Dr Takumi Takeuchi, ttakeuch@me.com

Received 9 December 2009; Revised 10 March 2010; Accepted 10 May 2010; Published online 21 June 2010.


Appropriate semen processing and assessment are critical for successful infertility treatment. We investigated whether laboratory procedures including semen preparation and incubation affect sperm DNA integrity. A total of 153 infertile men were involved. Conventional semen parameters and sperm chromatin structure assay (SCSA) parameters, that is, DNA fragmentation index (%DFI) and high DNA stainability (%HDS), were assessed on the fresh ejaculated semen samples, which were treated and incubated under different conditions. Negative correlations were identified between the %DFI and sperm concentration, motility, progressive motility and morphology. A lower percentage of DFI was detected in spermatozoa when density gradient centrifugation (DGC) was followed by swim-up treatment in comparison with DGC alone (P < 0.01). Although the %DFI increased in a time-dependent manner with incubation both at room temperature (RT) and at 37°C in air, the %DFI after 24 h at RT was significantly lower than that at 37°C (P < 0.05). Incubation with 5% CO2 was effective in maintaining sperm motility (P < 0.01); however, it induced further elevation of %DFI (P < 0.001). Thus, sperm DNA damage was associated with longer incubation periods. Interestingly, common culture conditions, such as maintaining pH and temperature, compromised the sperm DNA integrity.


density gradient centrifugation; DNA damage; male infertility; sperm chromatin structure assay; spermatozoa

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