Peterson J D, Miller S D, Waltenbaugh C
Department of Microbiology-Immunology, Northwestern University Medical School, Chicago, IL 60611.
J Virol Methods. 1990 Feb;27(2):189-201. doi: 10.1016/0166-0934(90)90135-3.
A rapid and efficient method is described for isotype quantitation of antiviral antibodies in mice infected with Theiler's murine encephalomyelitis virus (TMEV). Serum antibodies were reacted with fluorochrome-labeled TMEV in a modified fluid-phase particle concentration fluorescence immunoassay (PCFIA). Biotin and avidin were used to attach anti-immunoglobulin isotype antibodies to polystyrene particles by the separate incubation of biotinylated goat anti-mouse isotypes (IgG1-, IgG2a-, IgG2b-, IgG3-, or IgM-specific) with avidin coupled polystyrene particles. These anti-isotype particles captured the virus-antibody complexes. Mouse myeloma proteins were used to quantitate and standardize isotype profiles of normal mouse serum using fluorescein isothiocyanate (FITC)-labeled, goat anti-mouse isotypes and polystyrene particles coated with goat anti-mouse. These assays quantitated the affinity-purified mouse serum antiviral antibodies for the standardization of antiviral isotype assays. Immunoglobulin of all serum isotypes as well as the amount of virus-specific isotypes can be quantitated rapidly and accurately.
描述了一种快速有效的方法，用于对感染泰勒氏鼠脑脊髓炎病毒（TMEV）的小鼠体内抗病毒抗体进行亚型定量。在改良的液相颗粒浓度荧光免疫分析（PCFIA）中，血清抗体与荧光染料标记的TMEV反应。通过将生物素化的山羊抗小鼠亚型（IgG1、IgG2a、IgG2b、IgG3或IgM特异性）与抗生物素蛋白偶联的聚苯乙烯颗粒分别孵育，使用生物素和抗生物素蛋白将抗免疫球蛋白亚型抗体连接到聚苯乙烯颗粒上。这些抗亚型颗粒捕获病毒-抗体复合物。使用异硫氰酸荧光素（FITC）标记的山羊抗小鼠亚型和包被有山羊抗小鼠的聚苯乙烯颗粒，用小鼠骨髓瘤蛋白对正常小鼠血清的亚型谱进行定量和标准化。这些分析对抗病毒亚型分析的标准化定量了亲和纯化的小鼠血清抗病毒抗体。所有血清亚型的免疫球蛋白以及病毒特异性亚型的量都可以快速准确地定量。
