Ijaz A, Hunter A G
Department of Animal Science, University of Minnesota, St. Paul 55108.
J Dairy Sci. 1992 Feb;75(2):394-8. doi: 10.3168/jds.S0022-0302(92)77774-1.
Bovine sperm acquire the ability to penetrate zona-free hamster oocytes (capacitation) after incubation in TEST-yolk buffer. Our objective was to determine whether such sperm could penetrate zona-intact bovine oocytes in vitro. Bovine cumulus enclosed oocytes from 2- to 5-mm follicles were incubated in maturation medium for 24 h at 37 degrees C. Ejaculated bovine semen was diluted 1: 10 in TEST-yolk buffer, cooled to 4 degrees C, and stored for 8 h to induce capacitation. Sperm were then washed thrice in pH 7.6, .15 M NaCl containing .1% bovine serum albumin V (37 degrees C) and diluted to 2 x 10(6) sperm/ml in fertilization medium. Droplets of fertilization medium containing capacitated sperm, killed sperm, or no sperm were made under paraffin oil. Oocytes (matured 24 h) were added and cocultured with sperm for 8 h and then transferred to fresh fertilization medium for 40 h. After 24 h, 53% of the oocytes had matured (metaphase II). The fertilization rate of the metaphase II oocytes (203) with TEST-yolk capacitated sperm was 87%, whereas the parthenogenetic controls were 2 and 0%, respectively. Therefore, TEST-yolk buffer can be used to capacitate bull sperm for in vitro fertilization.
牛精子在TEST-卵黄缓冲液中孵育后获得穿透去透明带仓鼠卵母细胞的能力（获能）。我们的目的是确定这种精子是否能在体外穿透完整透明带的牛卵母细胞。将来自2至5毫米卵泡的牛卵丘-卵母细胞复合体在成熟培养基中于37℃孵育24小时。将射出的牛精液在TEST-卵黄缓冲液中按1:10稀释，冷却至4℃，并储存8小时以诱导获能。然后将精子在pH 7.6、含0.1%牛血清白蛋白V的0.15 M氯化钠溶液（37℃）中洗涤三次，并在受精培养基中稀释至2×10⁶精子/毫升。在石蜡油下制备含有获能精子、失活精子或无精子的受精培养基液滴。加入卵母细胞（成熟24小时）并与精子共培养8小时，然后转移到新鲜的受精培养基中培养40小时。24小时后，53%的卵母细胞成熟（处于中期II）。用TEST-卵黄获能精子处理的中期II卵母细胞（203个）的受精率为87%，而孤雌生殖对照组的受精率分别为2%和0%。因此，TEST-卵黄缓冲液可用于使公牛精子获能以进行体外受精。
