Evaluation of the Interaction between Exposure Time and Cryoprotectants Concentration on the Vitrification of Bovine Immature Oocytes with Cryotop

Document Type : Original Article

Authors
Kaveh Alizadeh 1
Najmeh Davoodian
Ali Kadivar 2
Ebrahim Ahmadi 3
Naser Shams Esfandabadi 2
1 PhD Student in Veterinary Reproductive Technologies, Department of Clinical Sciences, Faculty of Veterinary Medicine, Shahrekord University, Shahrekord, Iran
2 Department of Clinical Sciences, Faculty of Veterinary Medicine, Shahrekord University, Shahrekord, Iran.
3 Research Institute of Animal Embryo Technology, Shahrekord University, Shahrekord, Iran.
10.22034/ijvcs.2026.14938.1094
Abstract
This study aimed to evaluate the interaction between exposure time and concentration of cryoprotective agents (CPAs) in the vitrification of bovine immature oocytes. A total of 212 cumulus-oocyte complexes (COCs) with optimal quality were subjected to five different vitrification protocols using the Cryotop method. Independent variables included various concentrations of ethylene glycol (EG) and dimethyl sulfoxide (DMSO) in equilibration (EM) and vitrification (VM) media, along with different exposure times (30 seconds to 3 minutes). Morphological assessment revealed that Protocol 1 (1 min EM with 10% EG/DMSO and 30 sec VM with 20% EG/DMSO) yielded the highest percentage of morphologically intact oocytes (83.34±4.24), though none achieved Grade C quality (live oocyte and <50%dead cumulus cells). In contrast, Protocol 4 (2 minutes EM with 10% EG/DMSO and 45 seconds VM with 20% EG/DMSO) demonstrated the highest Grade C (44.18±0.07). Logistic regression analysis confirmed that 30-second exposure in VM significantly increased the likelihood of Grade B (live oocyte and >50%dead cumulus cells) classification (P<0.001), whereas extending EM time to 2 minutes decreased the probability of Grade C status (P<0.001). These findings indicated that successful vitrification of bovine immature oocytes depends on the optimal balance between CPA concentration and exposure time, with Protocol 4 recommended as the optimal method for simultaneous preservation of morphology and viability.
Keywords
Vitrification
Bovine immature oocyte
Cryoprotectant agents
Viability
Subjects
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Iranian Journal of Veterinary Clinical Sciences
Volume 19, Issue 2 - Serial Number 33
Fall and Winter 2025/2026
March 2026
Pages 61-69