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20001
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Subject: Animal Reproduction and Gynecology × End date: 2009 × Start date: 2000 × Thesis Type: Ph.D ×
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    ThesisItemOpen Access
    MORPHOLOGY AND VIABILITY OF BOVINE EMBRYOS FROZEN IN MEDIA CONTAINING BSA AND PROPANEDIOL
    (COLLEGE OF VETERINARY AND ANIMAL SCIENCES Mannuthy - Thrissur, 2000-10) RAMACHANDRAN, K.; . Suresan Nair, S.P
    The observation of the study was to compare the effect of Bovine Serum Albumin (BSA) and 1,2 Propanediol on the morphology and viability of bovine embryos frozen under two freezing and thawing protocols, A total of sixteen crossbred cows, kept under identical conditions, maintained in the Network Project on Embryo Transfer attached to the Department of Animal Reproduction, College of Vetennary and Animal Sciences, Thrissur. The animals were superovulated by Folltropm-V and Prosolvm, starting on the day 11 of the cycle. Of the 16 cows superovxilated 13 showed good response. While two cows did not show any response, there were multiple follicles in both the ovanes without any evidence of ovulation in the third animal. A total of 85 (76 transferable and 9 non transferable) embryos were recovered from a total of 24 flushings from 13 cows, non-surgically on day 7. A total of 56 embryos (mean 3.50 ± 0.822) were recovered m the first treatment, from 13 flushings. In one cow, though, there was 80 per cent flushing, no embryos could be recovered. While 22 embryos (mean 2.75 ± 0.861) were recovered m the second treatment from 8 flushings, only 7 (mean 1.4 ± 0.4) embryos were recovered in the third treatment, from 5 flushings. A total of 72 transferable (60 morulae and 12 blastocysts) were sleeted for the freezing tnals. The embryos were divided into three groups with 24 (20 morulae and 4 blastocysts) embryos and assigned to three media. The first medium was FBS with !0 per cent glycerol, second PBS containing 10 per cent glycerol and 1 per cent BSA; the third medium was with a composition of 10% glycerol and 20% 1,2 propanediol in PBS. Two freezing protocols were used for freezing of the embryos. In the first protocol, with 12 embryos (10 morulae and 2 blastocysts), the initial cooling was at a rate of 1 °C/min from room temperature to -6°C and then at a rate of 0.3°C/min to -35°C, while in the second protocol the initial rate of cooling was at 5°C/mm to -7°C and then at 0.3°C/mm to -30°C before transferring to liquid nitrogen. Thawing was earned out at 37°C for 20 sec after 30 days of preservation. Cryoprotectants were removed by two methods, a four step-wise using decreasing concentrations of cryoprotectants m the first method and one step using 1M sucrose phosphate buffered saline in the second. Thirty four embryos (26 morulae and 8 blastocysts) found normal after freezing and thawing were subjected to culture for 24 h in PBS enriched with 4 per cent BSA at 37°C and 5 per cent CO2 tension. Sixteen embryos (13 morulae and 3 blastocysts) were transferred to 15 recipient cows. While one cow was confirmed pregnant on examination 60 days after transfer, eleven cows returned to heat subsequently, two cows came to oestrus on days 34 and 35 respectively, after the transfer. The third showed oestrum on 45*'' day of transfer. The glucose, acid phosphatase and alkaline phosphatase values showed a normal range of 86.2 to 195.2 mg/100 ml; 14.17 KA to 22.3 KA and 119.02 to 129.00 KA (mean 128.075 ± 9.019, 18.675 ± 0.667 and 122.67 ± 0.788) respectively in the lummal fluid of the recipient animals. The average serum progesterone levels on day 0, 14 and 28 days after oestrus in 11 cows which showed subsequent heat after the transfer were 0.357 ±2.140, 3.053 ± 0.420 and 2.572 ± 0.627 ng/ml and that of the animals which failed to show oestrum were 0.157 ± 0.166, 3.793 ± 0.406 and 3.867 ± 0.362 ng/ml respectively. While significant difference was seen between the freezing media I and II and II and III respectively on the morphology of embryo after the freezing and thawing, no significant differences were seen between the media I and III, between the freezing protocols and cryoprotectant removal procedures on the morphology of embryos frozen. No significant differences were noticed on the effect of the freezing media, freezing protocols and the cryoprotectant removal procedure after the culture on the morphology of the embryos.