DEVELOPMENT OF MULTIPLE-IMMUNOASSAY FOR BIOACTIVE SUBSTANCES.

多重分析是指从单个样品中同时测量多种生物活性物质，可以从最少量的生物材料中获得许多信息。我们首先在放射免疫测定法（RIA）中工作，并成功地使用第一个抗体包被的孔连续测量激素。从保存宝贵的第一抗体的角度出发，我们利用第二抗体包被的孔进行研究，建立了大鼠垂体前叶激素的串联RIA。然后，为了建立平行分析，我们用第二抗体包被聚苯乙烯珠，然后用第一抗体处理它们。我们发现1/2英寸的珠粒在串联分析中表现优异，但1/4英寸珠粒在平行分析中表现更好。最终建立了多重ria协议。使用NIDDK试剂盒对大鼠垂体激素组合进行平行测定，结果表明，由于NIDDK TSH标准制剂中LH受到污染，LH不应与TSH同时测定，而其他组合则没有任何麻烦。我们进一步研究了使用eu标记的第一抗体开发多重TR-IFMA（时间分辨免疫荧光测定）。至此，我们成功建立了两栖动物促性腺激素、大鼠LH、大鼠PRL、大鼠、猪、人胰岛素、猪FSH和LH的单TR-IFMA。我们现在正致力于将这些分析方法应用于多种分析。

Multiple assay, which measures simultaneously many bioactive substances from a single sample, makes it possible to obtain many information from the least amount of biological materials. We first worked in RIA (radioimmunoassay) , and were successful in measuring hormones serially using first antibody-coated wells. From the standpoint to save the precious first antibodies, we then studied using wells coated with the second antibody, and establidhed serial RIA for rat anterior pituitary hormones. Then, in order to establish parallel assays, we coated polystirene beads with the second antibody, then treated them with the first antibody. We found that 1/2 inch beads were excellent in serial assay, but that 1/4 inch beads were better in parallel assays. Multiple-RIA protocol was finally established. Examination for the combination of rat pituitary hormones for parallel assays using NIDDK kits indicated that LH should not be measured simultaneously with TSH because of the contamination of LH in NIDDK TSH standard preparation, and that other combinations did not make any trouble.We further studied to develop the multiple TR-IFMA (time-resolved immunofluorometric assay) using Eu-labelled first antibody. So far, we succeeded in establishing single TR-IFMA for amphibian gonadotropin, rat LH,rat PRL,rat, porcine, human insulin, porcine FSH and LH.We are now engaged in applying these assay methods to multiple assay.