Biotin IgM Antibodies in Human Blood: A Previously Unknown Factor Eliciting False Results in Biotinylation-Based Immunoassays
Biotin IgM Antibodies in Human Blood: A Previously Unknown Factor Eliciting False Results in Biotinylation-Based Immunoassays
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DOI：
10.1371/journal.pone.0042376
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发表时间：
2012-08-03
期刊：
影响因子：
3.7
通讯作者：
Hedman, Klaus
中科院分区：
文献类型：
--
作者：
Chen, Tingting;Hedman, Lea;Hedman, Klaus
生物素是一种必需的维生素，其以高亲和力和特异性结合链霉亲和素或亲和素。由于生物素是一种可以与蛋白质连接而不影响其生物活性的小分子，因此生物素化被广泛应用于生物化学分析中。本实验室建立了以辣根过氧化物酶标记的链霉亲和素检测生物素化病毒样颗粒（VLP）为抗原的μ-捕获IgM酶免疫分析（EIA）。我们最近遇到一个血清样品与生物素化的VLP反应，但不与未生物素化的，这表明在人类血清中的生物素反应性抗体的发生。在本研究中，我们搜索了一般人群（612例成人血清样本和678例儿童血清样本）的IgM抗体与生物素反应，并制定了间接EIA定量的水平和评估其血清阳性率。这些IgM抗体存在于3%的成人中，无论年龄大小，但很少在儿童中发现。评估了生物素IgM对基于生物素化的免疫测定的不良影响，包括4种内部和1种商业病毒IgM EIA，表明生物素IgM确实会导致假阳性。生物素不能同时结合IgM和链霉亲和素或亲和素，表明这些生物素相互作用的化合物竞争共同的结合位点。在竞争性抑制试验中，生物素IgM抗体的亲和力范围为2.1 × 10 ~（-3）~ 1.7 × 10 ~（-4）mol/L。这是第一份关于在人体中发现的生物素抗体的报告，提供了关于基于生物素化的免疫测定的新信息，以及对维生素生物医学效应的新见解。
Biotin is an essential vitamin that binds streptavidin or avidin with high affinity and specificity. As biotin is a small molecule that can be linked to proteins without affecting their biological activity, biotinylation is applied widely in biochemical assays. In our laboratory, IgM enzyme immuno assays (EIAs) of mu-capture format have been set up against many viruses, using as antigen biotinylated virus like particles (VLPs) detected by horseradish peroxidase-conjugated streptavidin. We recently encountered one serum sample reacting with the biotinylated VLP but not with the unbiotinylated one, suggesting in human sera the occurrence of biotin-reactive antibodies. In the present study, we search the general population (612 serum samples from adults and 678 from children) for IgM antibodies reactive with biotin and develop an indirect EIA for quantification of their levels and assessment of their seroprevalence. These IgM antibodies were present in 3% adults regardless of age, but were rarely found in children. The adverse effects of the biotin IgM on biotinylation-based immunoassays were assessed, including four inhouse and one commercial virus IgM EIAs, showing that biotin IgM do cause false positivities. The biotin can not bind IgM and streptavidin or avidin simultaneously, suggesting that these biotin-interactive compounds compete for the common binding site. In competitive inhibition assays, the affinities of biotin IgM antibodies ranged from 2.1x10(-3) to 1.7x10(-4) mol/L. This is the first report on biotin antibodies found in humans, providing new information on biotinylation-based immunoassays as well as new insights into the biomedical effects of vitamins.