Comparison of various methods of processing human cryopreserved-thawed semen samples

Author： Srisombut C Morsheda M Lin MH Nassar A Oehninger S

ISSN： 1460-2350

Source： Human Reproduction, Vol.13, Iss.8, 1998-08, pp. : 2151-2157

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Abstract

We compared the efficacy of various methods of processing cryopreserved-thawed samples for the recovery of functionally adequate spermatozoa as assessed by the response to the sperm stress test (SST), an index of temperature activated sperm membrane lipid peroxidation, and immediate and delayed changes in sperm viability and motion parameters. Donor semen samples (n = 28) were cryopreserved-thawed and divided into six equal parts, one part was used as control and the remaining parts were used to compare five methods of sperm processing as follows: direct Percoll gradient processing, washing by one-step or stepwise addition of the washing medium followed by Percoll processing, and washing by one-step or stepwise addition of the washing medium. Additional samples (n = 10) were evaluated for the immediate and delayed (6 h at 37°C) impact of one-step and stepwise washing (without Percoll separation). Compared with wash-only methods, samples processed using Percoll had a significantly higher SST score (P = 0.001), motility, rapid spermatozoa (>50 μm/s), curvilinear velocity and motility index (P < 0.001).="" comparing="" various="" percoll="" methods,="" direct="" percoll="" processing="" resulted="" in="" the="" highest="" number="" of="" motile="" spermatozoa="" recovered="">P < 0.00001)="" and="" a="" higher="" sst="" score="" based="" on="" curvilinear="" velocity="">P = 0.001). Stepwise washing gave a significantly higher number of motile spermatozoa (P < 0.001)="" but="" with="" a="" significantly="" lower="" sst="" score="" based="" on="" the="" concentration="" of="" motile="" spermatozoa="">P = 0.001), motility (P = 0.001) and motility index (P = 0.01). Sperm viability and motion parameters after 6 h of incubation showed no difference between one-step and stepwise washing. In conclusion, compared with wash-only methods, Percoll processed samples resulted in the recovery of spermatozoa with superior quality as assessed by SST and motion analysis. One-step washing of the samples gave an overall comparable recovery compared to the samples prepared stepwise. Having significantly higher SST scores, similar viability and the maintenance of motility, one-step washing may be a better method of processing thawed samples than the stepwise washing. Keywords:cryopreservation/gradient separation/human spermatozoa/sperm stress test/sperm washing