EFFECT OF DIFFERENT CATEGORIES OF SERA AND BOVINE SERUM ALBUMIN ON IN VITRO MATURATION OF SURTI BUFFALO OOCYTES

EFFECT OF DIFFERENT CATEGORIES OF SERA AND BOVINE SERUM ALBUMIN ON IN VITRO MATURATION OF SURTI BUFFALO OOCYTES Student: Chirag N. Mistry Major Advisor: Dr. A.J. Dhami Department of Animal Biotechnology College of Veterinary Science and Animal Husbandry Anand Agricultural University, Anand 388 001 This study was conducted over a period of 6 months from September 2008 to February 2009 with the objectives of evaluating the effects of different concentrations (0.05, 0.1, 0.3, 0.6 and 0.9 per cent) of BSA-FAF and also of different categories of sera like Fetal Calf Serum (Gibco), Fetal Buffalo Serum, Oestrus Buffalo Serum, Postoestrus Buffalo Serum and Anestrus Buffalo Serum (all @ 20%) in relation to standard BSA (0.6%) on in vitro maturation of buffalo oocytes in TCM-199 medium. The rate of maturation was confirmed both by cytoplasmic and nuclear maturation using Hoechst stain 33342. A total of 456 ovaries of Surti buffaloes collected from the local slaughter house were transported to the laboratory at 30°C in normal saline solution within I72 hours of slaughter of animal for further processing. In all 1409 oocytes were recovered fromthem by using slicing method of surface follicles. The sera samples used for culture were obtained from the animals showing different stages of oestrous cycle and were heat inactivated in the laboratory. An average number of follicle of small, medium and large size found per ovary was 0.82, 0.48 and 0.24, respectively, with an overall mean of 1.55. The distribution of these follicles came to 53.18, 31.12-and 15.70 percent, respectively. The slicing method of oocyte recovery gave quite good result. The average oocyte recovery was 3.09 per ovary. The average recovery rate of Grade A, Grade B and Grade C oocyte was 1.02,1.22 and 0.85, respectively. The maturation rate with 0.05, 0.1, 0.3, 0.6 and 0.9 per cent concentration of BSA in TCM-199 was found to be 44.52, 50.99, 59.02, 84.43 and 64.29 per cent, respectively. The BSA 0.6 per cent yielded the highest maturation rate, which differed significantly from other BSA concentrations. The maturation rate for locally prepared FOBS, AnBS, FCS (Gibco), FBS and OBS was found to be 64.63, 54.55, 70.63, 60.48 and 78.16 per cent, respectively. The medium containing oestrus buffalo serum yielded significantly higher (P<0.05) maturation rate than the others, though it was little less than the BSA 0.6. The highest maturation of Grade A oocytes was found in BSA 0.6 followed by OBS and other sera. While in Grade B it was in BSA 0.9 followed by BSA 0.6, and for Grade C oocytes the highest maturation rate was with BSA 0.6 followed by FOBS. According to nuclear maturation, the highest number of oocytes with germinal vesicle was found in medium containing FBS (25.21 %) followed by AnBS and others. The highest number of germinal vesicle break down was found in FOBS (30.61 %) followed by FCS (Gibco) and others. The higher number of oocytes with Metaphase-I was in the medium containing BSA 0.6 per cent (22.13 %) followed by FCS (Gibco), while, the Metaphase-II stage was found to be higher in medium containing OBS (41.38 %) followed by BSA 0.6 and others. Degenerated oocytes were maximum with BSA 0.05 per cent (40.41 %) and minimum with OBS (9.20 %). It was concluded that BSA concentration of 0.6 per cent is the optimum for in vitro maturation of buffalo oocytes, and that OBS can be used instead of BSA as a cheaper and easily available source of serum for in vitro maturation of buffalo oocytes.