Moncayo H, Moncayo R, Benz R, Wolf A, Lauritzen C
Department of Obstetrics and Gynecology, University of Ulm, Federal Republic of Germany.
J Clin Invest. 1989 Dec;84(6):1857-65. doi: 10.1172/JCI114372.
We developed an ELISA system for the detection of human anti-ovarian antibodies. Bovine corpora lutea were extracted in PBS (pH 7.2) and fractionated by ultracentrifugation. Both the soluble fraction obtained after 80,000 g (S80) and the Triton-extracted membrane fraction (ST288) were used as antigens. Additionally, the luteinizing hormone (LH)/human chorionic gonadotropin (hCG) receptor was isolated by affinity chromatography (wheat germ agglutinin and LH-Sepharose) and also used as an antigen. In 7 of 14 patients with primary sterility and endometriosis a positive reaction was observed. Similarly, 6 of 16 patients with secondary sterility and endometriosis were also positive. Patients being stimulated for in vitro fertilization and presenting either primary or secondary sterility were positive in 5 of 22 and 6 of 16 cases, respectively. In the S80 test 41 of 60 sera presented IgG2 antibodies, whereas in the ST288 test 38 of 60 belonged to the IgG1 subclass. Kappa and lambda chains were equally distributed. Some patients could recognize the unoccupied LH/hCG receptor as an antigen, while others recognized only the complex formed by the hormone plus the hormone receptor. The S80 and ST288 antigens were isolated by affinity chromatography. Gel permeation of the purified antigens revealed in each case the presence of an antigen complex. The apparent molecular weight was between 2,000 and 36,000 D. Cross-reactivity studies using affinity-purified antibodies demonstrated an antigenic relationship of the membrane, soluble, and extractable fractions. NAc-(beta-1----4)-D-glucosaminide and -D-galactopyranoside were the main terminal glycosides.
我们开发了一种用于检测人抗卵巢抗体的酶联免疫吸附测定(ELISA)系统。牛黄体在磷酸盐缓冲盐水(PBS,pH 7.2)中提取,然后通过超速离心进行分级分离。80,000g离心后获得的可溶性部分(S80)和经曲拉通提取的膜部分(ST288)均用作抗原。此外,通过亲和色谱法(麦胚凝集素和促黄体生成素-琼脂糖凝胶)分离出促黄体生成素(LH)/人绒毛膜促性腺激素(hCG)受体,并将其也用作抗原。在14例原发性不育和子宫内膜异位症患者中有7例观察到阳性反应。同样,16例继发性不育和子宫内膜异位症患者中有6例也呈阳性。接受体外受精刺激且患有原发性或继发性不育的患者中,分别有22例中的5例和16例中的6例呈阳性。在S80检测中,60份血清中有41份呈现IgG2抗体,而在ST288检测中,60份中有38份属于IgG1亚类。κ链和λ链分布均等。一些患者能够将未被占据的LH/hCG受体识别为抗原,而另一些患者仅识别由激素加激素受体形成的复合物。S80和ST288抗原通过亲和色谱法分离。对纯化抗原进行凝胶渗透分析,在每种情况下均显示存在抗原复合物。表观分子量在两千至36,000道尔顿之间。使用亲和纯化抗体进行的交叉反应研究表明,膜部分、可溶性部分和可提取部分之间存在抗原关系。N-乙酰基-(β-1→4)-D-葡糖胺和-D-吡喃半乳糖苷是主要的末端糖苷。