Effect of cryoprotectants and their concentration on in vitro development of vitrified-warmed immature oocytes in buffalo (Bubalus bubalis).
Experiments were conducted to study the effect of cryoprotectants, dimethyl sulfoxide (DMSO), ethylene glycol (EG), 1,2-propanediol (PROH), and glycerol at different concentrations (3.5, 4, 5, 6, and 7 M each with 0.5 M sucrose and 0.4% BSA in DPBS) on survival, in vitro maturation, in vitro fertilization, and post-fertilization development of vitrified-thawed immature buffalo oocytes. The COCs were harvested from the ovaries by aspirating the visible follicles. The recovery of post-thaw morphologically normal oocytes was lower in 3.5 and 4 M DMSO, EG, and PROH compared to 5, 6, and 7 M. In all the concentrations of glycerol, an overall lower numbers of oocytes recovered were normal compared to other cryoprotectants. Less number of oocytes reached metaphase-II (M-II) stage from the oocytes cryopreserved in any of the concentrations of DMSO, EG, PROH, and glycerol compared to fresh oocytes. Among the vitrified groups, highest maturation was obtained in 7 M solutions of all the cryoprotectants. The cleavage rates of oocytes vitrified in different concentrations of DMSO, EG, PROH, and glycerol were lower than that of the fresh oocytes. The cleavage rates were higher in oocytes cryopreserved in 6 and 7 M DMSO, EG, PROH, and glycerol compared with oocytes cryopreserved in other concentrations. However, the percentage of morula and blastocyst formation from the cleaved embryos did not vary in fresh oocytes and vitrified oocytes. In conclusion, this report describes the first successful production of buffalo blastocysts from immature oocytes cryopreserved by vitrification.
Wani
N A
NA
Embryo Biotechnology Laboratory, Department of Animal Reproduction, Gynaecology and Obstetrics, College of Veterinary Sciences, G. B. Pant University of Agriculture and Technology, Pantnagar, India. dr_nawani@yahoo.com.in
Maurya
S N
SN
Misra
A K
AK
Saxena
V B
VB
Lakhchaura
B D
BD
D000818:Animals; D001755:Blastocyst / Q000502:physiology; D002020:Buffaloes; D002970:Cleavage Stage, Ovum / Q000187:drug effects; D015925:Cryopreservation / Q000662:veterinary*; D003451:Cryoprotective Agents / Q000008:administration & dosage* / Q000633:toxicity; D046508:Culture Techniques; D004121:Dimethyl Sulfoxide / Q000008:administration & dosage; D005314:Embryonic and Fetal Development; D019855:Ethylene Glycol / Q000008:administration & dosage; D005260:Female; D005307:Fertilization in Vitro / Q000662:veterinary*; D005990:Glycerol / Q000008:administration & dosage; D006358:Hot Temperature; D009028:Morula / Q000502:physiology; D009865:Oocytes / Q000187:drug effects / Q000502:physiology*; D019946:Propylene Glycol / Q000008:administration & dosage; D012710:Serum Albumin, Bovine / Q000008:administration & dosage; D013395:Sucrose / Q000008:administration & dosage
D003451:Cryoprotective Agents; D012710:Serum Albumin, Bovine; D013395:Sucrose; D019946:Propylene Glycol; D019855:Ethylene Glycol; D005990:Glycerol; D004121:Dimethyl Sulfoxide