In vitro fertilization of bovine oocytes by spermatozoa capacitated in vitro.

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In vitro fertilization of ovulated bovine oocytes was attempted with fresh or frozen semen after capacitation in vitro. Sperm incubations and oocyte cultures were performed in a Krebs Ringer bicarbonate medium containing Na pyruvate, glucose and .3% bovine serum albumin. High ionic strength (HIS) medium was prepared by adding NaCl to provide an osmolarity of 370 to 380 nOsmol/kg. Donor cows were treated with prostaglandin F2 alpha and a series of follicle-stimulating hormone injections and ovulated oocytes were recovered 72 h after prostaglandin treatment. Fresh or frozen semen was: 1) placed directly into a microdrop of standard medium (SM) under oil; 2) washed by centrifugation with SM and placed in a microdrop of SM or 3) pretreated with HIS for 10 min, washed and placed into a microdrop of SM. In all cases, spermatozoa were preincubated for 3 h at a concentration of approximately 10(6) cells/ml before addition of oocytes. Oocytes were incubated with spermatozoa for 6 h, transferred to fresh medium and cultured for 24 h. When spermatozoa were placed directly into a microdrop, three of 34 (9%) oocytes were penetrated, but none divided. Spermatozoa washed with SM penetrated 20 of 45 (44%) oocytes and three (7%) divided. Spermatozoa incubated in HIS penetrated 14 of 47 (30%) oocytes and five (11%) divided. The washing of spermatozoa with standard medium was equally as effective as incubation with high ionic strength medium in inducing in vitro capacitation of bovine spermatozoa.

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Journal of animal science

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