Cryoprotectants addition and removal are essential steps during oocytes cryopreservation

however

high concentration of cryoprotectants may cause osmotic and toxic injuries to oocytes. In order to study the injuries of porcine MII-stage oocytes during cryoprotectants addition and removal processes

cryoprotectant was loaded and unloaded to/from oocytes with traditional methods (one-step method

two-step method)

the effects of vitrification solution

thawing solution and dilution solution on the survival rate of oocytes were investigated. The experimental results showed that the osmotic injuries to oocytes happened mainly in two stages: the transfer process from vitrification solution to thawing solution

and the cryoprotectants removal process. The concentration of thawing solution and equilibrium time had a great influence on the survival rate of oocytes. The cryoprotectants removal process was highly correlated with addition process

and the selection of thawing solution concentration and equilibrium time relied on the vitrification solution concentration and equilibrium time. These results could provide a reference for cryopreservation protocols of porcine MII-stage oocytes.