Aseptic Cryoprotectant-Free Vitrification of Human Spermatozoa by Direct Dropping into a Cooling Agent

Spermatozoa cryopreservation is used for the management of infertility and some other medical conditions. Routinely applied cryopreservation techniques depend on permeating cryoprotectants and relatively slow freezing rates. Cryoprotectant-free vitrification is an alternative and cost-effective method that is based on rapid cooling of spermatozoa by direct plunging into a cooling agent to prevent lethal intracellular ice crystallization and the detrimental effects of high salt concentrations. One of the problems with this technique is that full sterilization of commercially produced liquid nitrogen, which could be contaminated with different pathogens, is not possible. Here we use a benchtop device for the production of sterile liquid air with the same temperature as liquid nitrogen (-195.7 °C). This has been used to develop aseptic technology for cryoprotectant-free vitrification of human spermatozoa.