Mouse sperm antigens that participate in fertilization. I. Inhibition of sperm fusion with the egg plasma membrane using monoclonal antibodies.
Mouse sperm antigens that participate in fertilization. I. Inhibition of sperm fusion with the egg plasma membrane using monoclonal antibodies.
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参与受精的小鼠精子抗原。
DOI：
10.1095/biolreprod33.2.515
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发表时间：
1985
影响因子：
3.6
通讯作者：
Waibel,R
中科院分区：
文献类型：
--
作者：
Saling,PM;Irons,G;Waibel,R
单克隆抗体（mab）已经产生，以确定负责与卵子相互作用导致受精的精子成分。在这里，我们报告了一组六个不同的单克隆抗体，它们都定位于精子头部的一个有限区域，赤道段。其中一些单克隆抗体显示出与其他被测物种（人、仓鼠、兔）精子的交叉反应性；当发生交叉反应时，尽管该同源区域在不同物种中具有不同的构型，但mAb的分布仅限于赤道段。当测试对体外受精过程的影响时，腹水中含有六种单克隆抗体中的两种，M29和M37，显示出显著的抑制作用。使用纯化的M29免疫球蛋白M观察这种抑制的浓度依赖性，在0至0.2 mg/ml的范围内。单抗对受精的抑制与卵周围的细胞层（积云）或非细胞层（透明带）的存在无关，表明这两种抗精子单抗的特异性抑制位点都是卵质膜。在12%十二烷基硫酸钠-聚丙烯酰胺凝胶上电泳分离精子成分，然后转移到硝化纤维素片上进行免疫检测，鉴定了该组中两个单克隆抗体识别的精子成分：抑制受精的M29和不抑制受精的M2。使用M29 mAb，检测到单个精子组分的表观亚基分子量约为40,000，而在用M2 mAb孵育的硝化纤维素条中，两个组分显示出反应性，一个非常突出的条带约为44,000，一个紧密的条带约为36,000。小鼠肝脏的平行硝化纤维素条没有显示出这些反应性，这与间接免疫荧光数据一致，在间接免疫荧光数据中，只有睾丸和精子，而没有肝、肾、卵巢和附睾上皮显示出阳性反应性。这些结果表明，使用单克隆抗体可以识别参与受精过程中单个事件的精子成分。此外，使用这种策略，我们已经确定了一种特定的精子成分，它似乎是精子与卵质膜融合的候选成分。
Monoclonal antibodies (mAbs) have been generated to determine the sperm components responsible for interaction with an egg that results in fertilization. Here, we report upon a group of six different mAbs, all of which localize to a restricted region of the sperm head, the equatorial segment. Several of these mAbs demonstrated cross-reactivity with sperm from the other species tested (human, hamster, rabbit); when cross-reaction occurred, the mAb distribution was restricted to the equatorial segment despite the various configurations that this homologous region assumes in different species. When tested for an effect upon the fertilization process in vitro, ascites fluids containing two of the six mAbs, M29 and M37, displayed significant inhibition. The concentration dependency of this inhibition was observed using purified M29 immunoglobulin M, over a range of 0 to 0.2 mg/ml. The mAb inhibition of fertilization was independent of the presence of either the cellular (the cumulus) or acellular (the zona pellucida) layers surrounding the egg, indicating that the specific locus of inhibition for both of these antisperni mAbs was the egg plasma membrane. Immunologic detection of sperm components separated by electrophoresis on 12% sodium dodecyl sulfate-polyacrylamide gels followed by transfer to nitrocellulose sheets was used to identify the sperm components recognized by two of the mAbs in this group: M29, which inhibited fertilization, and M2, which did not inhibit fertilization. Using M29 mAb, a single sperm component with an apparent subunit molecular weight of approximately 40,000 was detected, whereas in the nitrocellulose strips incubated with M2 mAb two components displayed reactivity, a very prominent band at approximately 44,000 and a tight cluster of bands at approximately 36,000. Parallel nitrocellulose strips of mouse liver did not display these reactivities, consistent with indirect immunofluorescence data in which only testis and sperm, and not liver, kidney, ovary, and epididymal epithelium, demonstrated positive reactivity. These results indicate that the use of mAbs permits identification of sperm components that participate, putatively, in individual events of the fertilization process. Furthermore, using this strategy, we have identified a specific sperm component that appears to be a candidate for a role in sperm fusion with the egg plasma membrane.