Preliminary Study On In Vitro Fertilization With Sorted Buffalo Sperm

The objective of the present studies was to test the feasibility of flow-cytometrically sorted buffalo sperm used in IVF systems and to establish suitable IVF systems for sorted buffalo sperm. Oocytes were aspirated from 2-6 mm follicles on the buffalo ovaries from slaughterhouse and matured in vitro. After maturation, oocytes were fertilized with sorted sperm for 8-10 h and then the presumptive zygotes were cultured with granulose cells. Cleavage and blastocyst rates per oocytes insemination were recorded on D2 and D6-D9 after insemination, respectively. Embryos were freshly transplanted to recipient buffalos or cryopreserved.There were significant difference in cleavage rate (42.23% vs 52.28%) and blastocyst rate (19.98% vs 28.59% ) between sorted and unsorted sperm(P<0.01); There were no significant difference in the proportions of blastocyst development on D6, D7 and D8 after insemination with sorted and unsorted sperm(P>0.05); The proportions of blastocyst development on D6, D7 and D8 after insemination with X-sperm and Y-sperm also showed no significant difference(P>0.05); Sorted sperm were stored at 4℃ for 1)15~18 2)39-42 and 3) 69-72 h after sorting, then fertilized with oocytes. The cleavage rate (39.57% vs 37.14%) or the blastocyst development rate (20.43% vs 18.29%) showed no significant difference between group 15 - 18 h and group 39 - 42 h , but significant difference between group 69-72 h and the other two groups (P<0.01) . Survival rate of post frozen-thawed embryos showed no difference between embryos produced by sorted sperm and unsorted sperm (70.93% vs 73.2%, P>0.05).The results above indicate that 1) Sorted buffalo sperm can be successfully used in IVF systems and produced sex controlled embryos for ET; 2) The blastocyst development was not delayed by sorting procedure; 3)Male and female embryos produced by IVF sorted sperm showed similar development speed 5) Embryos produced by IVF with sorted sperm can be successfullycryopreserved. 6) Pregnancy can be successfully established after ET with embryos produced from IVF with sorted buffalo sperm in these IVF systems.