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使用合成肽Env gp46(188 - 209)作为抗原,对血清中(抗人类I型T细胞白血病病毒)IgG进行免疫复合物转移酶免疫测定。

Immune complex transfer enzyme immunoassay for (anti-human T-cell leukemia virus type I) IgG in serum using a synthetic peptide, Env gp46(188-209), as antigen.

作者信息

Kohno T, Sakoda I, Ishikawa E

机构信息

Department of Biochemistry, Medical College of Miyazaki, Japan.

出版信息

J Clin Lab Anal. 1991;5(1):25-37. doi: 10.1002/jcla.1860050106.

Abstract

An immune complex transfer enzyme immunoassay for (anti-human T-cell leukemia virus type I) IgG (anti-HTLV-I IgG) in serum using a chemically and safely synthesized peptide, env gp46(188-209), is described. Anti-HTLV-I IgG in test serum, which had been incubated with excess of inactive beta-D-galactosidase to eliminate interference by anti-beta-D-galactosidase antibodies, was reacted simultaneously with dinitrophenyl bovine serum albumin-env gp46(188-209) conjugate and env gp46(188-209)-beta-D-galactosidase conjugate. The complex formed of the three components was trapped onto polystyrene balls coated with affinity-purified (anti-dinitrophenyl group) IgG. After washing to eliminate nonspecific IgG in the test serum and excess of the beta-D-galactosidase conjugate, the complex was eluted from the polystyrene balls with dinitrophenyl-L-lysine and transferred to polystyrene balls coated with affinity-purified (anti-human IgG gamma-chain) IgG. beta-D-Galactosidase activity bound to the (anti-human IgG gamma-chain) IgG-coated polystyrene balls was assayed by fluorometry. This assay was sensitive and detected anti-HTLV-I IgG in serum samples which were negative by the conventional enzyme immunoassay and Western blotting. And the specificity of this assay was confirmed by preincubation of test serum with excess of env gp46(188-209). However, some disadvantages were also noted.

摘要

本文描述了一种使用化学合成且安全的肽env gp46(188 - 209)检测血清中抗人T细胞白血病病毒I型IgG(抗HTLV - I IgG)的免疫复合物转移酶免疫测定法。将检测血清中的抗HTLV - I IgG与过量的无活性β - D - 半乳糖苷酶孵育以消除抗β - D - 半乳糖苷酶抗体的干扰后,使其与二硝基苯基牛血清白蛋白 - env gp46(188 - 209)偶联物和env gp46(188 - 209) - β - D - 半乳糖苷酶偶联物同时反应。由这三种成分形成的复合物被捕获到包被有亲和纯化的(抗二硝基苯基)IgG的聚苯乙烯球上。洗涤以去除检测血清中的非特异性IgG和过量的β - D - 半乳糖苷酶偶联物后,用二硝基苯基 - L - 赖氨酸从聚苯乙烯球上洗脱复合物,并转移到包被有亲和纯化的(抗人IgGγ链)IgG的聚苯乙烯球上。通过荧光法测定与(抗人IgGγ链)IgG包被的聚苯乙烯球结合的β - D - 半乳糖苷酶活性。该测定法灵敏,能检测出常规酶免疫测定和蛋白质印迹法呈阴性的血清样本中的抗HTLV - I IgG。并且通过用过量的env gp46(188 - 209)对检测血清进行预孵育,证实了该测定法的特异性。然而,也注意到了一些缺点。

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