Large volume cryoprotectant-free vitrification: an alternative to conventional cryopreservation for human spermatozoa

Vitrification is a simple and cost-effective method for the storage of human spermatozoa without the use of conventional cryoprotectants, by plunging the sperm suspension directly into liquid nitrogen. As a result, solidification of living cells without the formation of ice crystals is achieved during cooling. This study aimed to compare cryoprotectant-free vitrification to conventional cryopreservation protocols. Semen samples (n = 35) were collected from patients seeking diagnostic assistance at the Reproductive and Endocrine Unit at Steve Biko Academic Hospital. Samples were processed using a discontinuous density-gradient centrifugation method. Washed samples were split into two aliquots and cryopreserved either by means of cryoprotectant-free vitrification (sucrose + 1% albumin) or conventional slow freezing (TEST-yolk buffer). Post-thawing, the sperm motion parameters, mitochondrial membrane potential (Δψm) and DNA fragmentation were compared between the two groups. No significant differences were observed in the sperm motility parameters (P > 0.05). Significantly higher percentages of Δψm (11.99% ± 4.326% versus 6.58% ± 1.026%; P < 0.001) and lower percentages of DNA fragmentation (2.79% ± 1.017% versus 3.86% ± 1.38%; P < 0.01) were observed when comparing cryoprotectant-free vitrification to conventional cryopreservation. Cryoprotectant-free vitrification is a rapid and promising alternative to conventional methods resulting in good-quality spermatozoa post-thaw.