Vajta G, Peura T T, Holm P, Páldi A, Greve T, Trounson A O, Callesen H
Embryo Technology Center, Danish Institute of Agricultural Sciences, Research Centre Foulum, Tjele, Denmark.
Mol Reprod Dev. 2000 Mar;55(3):256-64. doi: 10.1002/(SICI)1098-2795(200003)55:3<256::AID-MRD3>3.0.CO;2-7.
Culture of mammalian zygotes individually and in small groups results in lower developmental rates than culture of large groups. Zona-free zygotes also have impaired developmental potential in current culture systems. This paper describes a new approach to resolve the problems, the Well of the Well (WOW) system. Small wells (WOWs) were formed in four-well dishes by melting the bottom with heated steel rods. The WOWs were then rinsed, the wells were filled with medium, and the embryos were placed into the WOWs. To test the value of the WOW system a 3 x 3 factorial experiment was performed. Bovine presumptive zygotes were cultured from day 1 to day 7 (day 0: day of insemination) using three modules (single embryos, embryo groups of five, or single zona-digested embryos) and three different culture systems (400 microl medium, 200 microl drops, or WOWs). An additional control group consisted of 40 to 50 embryos cultured in 400 microl medium. The WOW system resulted in higher blastocyst/oocyte rates for all three modules (single: 59%; group of five: 61%; single zona-digested: 53%) than the culture in drops or in wells (P < 0.05 for all). The developmental rate was independent of the number of WOWs per well. The cell number of blastocysts cultured in the WOW system did not differ from that of the controls. Apart from its theoretical value in revealing the role of different factors influencing embryo development in vitro, the WOW system may have immediate practical consequences in certain areas of mammalian embryo production.
与大群体培养相比，对哺乳动物合子进行单独或小群体培养会导致发育率降低。在当前的培养系统中，去除透明带的合子发育潜力也会受损。本文描述了一种解决这些问题的新方法——“井中井”（WOW）系统。通过用加热的钢棒熔化四孔培养皿的底部，在其中形成小井（WOW）。然后冲洗这些小井，向其中加入培养基，并将胚胎放入小井中。为了测试WOW系统的价值，进行了一项3×3析因实验。使用三个模块（单个胚胎、五个胚胎的组或单个消化透明带的胚胎）和三种不同的培养系统（400微升培养基、200微升液滴或WOW），将牛的推定合子从第1天培养到第7天（第0天：授精日）。另外一个对照组由在400微升培养基中培养40至50个胚胎组成。与在液滴或培养孔中培养相比（所有情况P<0.05），WOW系统在所有三个模块中都产生了更高的囊胚/卵母细胞率（单个胚胎：59%；五个胚胎的组：61%；单个消化透明带的胚胎：53%）。发育率与每个培养孔中小井（WOW）的数量无关。在WOW系统中培养的囊胚的细胞数量与对照组没有差异。除了在揭示影响体外胚胎发育的不同因素的作用方面具有理论价值外，WOW系统在哺乳动物胚胎生产的某些领域可能会立即产生实际影响。
