Comparison of Cryotop and micro volume air cooling methods for cryopreservation of bovine matured oocytes and blastocysts

Abstract

This study was designed to compare the efficiency of the Cryotop method and that of two methods that employ a micro volume air cooling (MVAC) device by analyzing the survival and development of bovine oocytes and blastocysts vitrified using each method. In experiment I, in vitro-matured (IVM) oocytes were vitrified using an MVAC device without direct contact with liquid nitrogen (LN2; MVAC group) or directly plunged into LN2 (MVAC in LN2 group). A third group of IVM oocytes was vitrified using a Cryotop device (Cryotop group). After warming, vitrified oocytes were fertilized in vitro. There were no significant differences in cleavage and blastocyst formation rates among the three vitrified groups, with the rates ranging from 53.1% to 56.6% and 20.0% to 25.5%, respectively; however, the rates were significantly lower (P < 0.05) than those of the fresh control group (89.3% and 43.3%, respectively) and the solution control group (87.3% and 42.0%, respectively). In experiment II, in vitro-produced (IVP) expanded blastocysts were vitrified using the MVAC, MVAC in LN2 and Cryotop methods, warmed and cultured for survival analysis and then compared with the solution control group. The rate of development of vitrified-warmed expanded blastocysts to the hatched blastocyst stage after 24 h of culture was lower in the MVAC in LN2 group than in the solution control group; however, after 48–72 h of culture, the rates did not significantly differ between the groups. These results indicate that the MVAC method without direct LN2 contact is as effective as the standard Cryotop method for vitrification of bovine IVM oocytes and IVP expanded blastocysts.

State of the Art in Fertility Preservation for Female Patients Prior to Oncologic Therapies

“…The Cryotop is considered to be one of the most efficient vitrification methods both for oocytes and embryos, providing high rates of survival for both humans and animal models, and like other open cryo-preservation systems, directly contacts liquid nitrogen, increasing the risk of viral contamination [40]. Alternative methods were devised, such as micro-volume air-cooling (MVAC) that focused on preventing direct contact of the specimen with the liquid nitrogen [41].…”

Fertility preservation during the COVID-19 pandemic: mitigating the viral contamination risk to reproductive cells in cryostorage

“…However, use of screw capped plastic vials can lead to contact with surrounding LN2 by vacuum creation in the vial due to condensation of the atmosphere at such low temperature thereby drawing in the liquid nitrogen ( Woods and Thirumala, 2011 ). Several closed carrier systems have been employed for cryopreservation, such as microvolume air cooling (MVAC) device ( Punyawai et al , 2015 ), high-security vitrification straw (HSV) or Cryotip, some of which have been used for human embryo cryopreservation thereby hermetically isolating the reproductive cells ( Arav, 2020 ; Abdel-Hafez et al , 2011 ; Kuwayama et al , 2005 ). Hence, ESHRE in its latest guidelines recommends the use of high-security straws and vapor-phase storage for cryopreservation of reproductive samples from COVID-19 positive patients ( ESHRE, 2020b ).…”

Features of the preparation of biological material for genome editing in cattle

“…It results in division of sperm into spermatozoids and semenal plasma by сentrifugalization in discontinuous Percoll gradient (Percoll 45 % and Percoll 90 %) for 5-10 minutes at 1500-3000 g and at room temperature. Sperm pellet after сentrifugalization is washed by buffer fluid for 3-5 minutes at 1000 g [23].…”

“…The process of vitrification starts with equilibration of oocytes/zygotes for 12-15 minutes at room temperature in the solution with low concentration of penetrating cryoprotectors, 3-4 % ethylene glycol or 7,5% ethylene glycol + 7,5 % dimethylsulfoxide. Af-ter equilibration oocytes/zygotes are placed in the medium for vitrification, containing 30-35 % ethylene glycol with addition of 1,0 mol/L saccharose or 15 % dimethylsulfoxide + 15 % ethylene glycol + 0.5 mol/L saccharose [14,23]. In the vitrification medium oocytes/zygotes should stay for not more than 1.5 minutes until their placing into liquid nitrogen, including the time of putting them on the device for vitrification.…”