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首页> 外文期刊>Journal of Clinical Microbiology >Enzyme-linked immunosorbent assay and enzyme-linked coagulation assay for detection of Clostridium botulinum neurotoxins A, B, and E and solution-phase complexes with dual-label antibodies.
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Enzyme-linked immunosorbent assay and enzyme-linked coagulation assay for detection of Clostridium botulinum neurotoxins A, B, and E and solution-phase complexes with dual-label antibodies.

机译：酶联免疫吸附测定和酶联凝血测定，用于检测肉毒梭菌神经毒素A，B和E以及具有双标记抗体的溶液相复合物。

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The measurement of toxins A, B, and E from Clostridium botulinum was accomplished by use of a modified sandwich enzyme-linked immunosorbent assay (ELISA) employing labeled horse antibody and either chicken antibody or biotinylated horse antibody. The complexes formed in solution phase were captured onto solid phases coated with rabbit anti-chicken immunoglobulin G (chicken antibody) or avidin (biotinylated antibody). The assay was brought to the sensitivity of the mouse bioassay (5 to 10 pg/ml, or 0.03 to 0.07 pM) by employing as labeling enzyme the factor X activator of Russell's viper venom (RVV-XA) and a sensitive coagulation-based assay amplification system known as enzyme-linked coagulation assay. Complex formation was found to be a slower reaction than binding to the capture plate, and so the assay used a preincubation step to produce the solution-phase complexes before they were bound to the solid phase. Keeping the concentrations of Russell's viper venom factor X activator antibody and capture antibody constant for diluted samples and diluting complexes into buffer without keeping labeled antibody concentrations constant were equivalent in allowing the detection of low neurotoxin concentrations. This ELISA-enzyme-linked coagulation assay procedure is a convenient alternative to the mouse bioassay, which shows complete resolution of the neurotoxins in addition to the requisite sensitivity.
机译：肉毒梭菌中毒素A，B和E的测定通过使用改良的夹心酶联免疫吸附测定（ELISA）进行，该测定采用标记的马抗体和鸡抗体或生物素化的马抗体。将溶液相中形成的复合物捕获到包被有兔抗鸡免疫球蛋白G（鸡抗体）或抗生物素蛋白（生物素化抗体）的固相上。通过将Russell毒蛇毒的X因子激活剂（RVV-XA）用作标记酶并进行基于灵敏的凝血的测定，将测定提高到小鼠生物测定的灵敏度（5至10 pg / ml，或0.03至0.07 pM）扩增系统称为酶联凝血测定。发现复合物的形成比与捕获板的结合反应慢，因此该测定法在结合固相之前使用了预温育步骤以生成溶液相复合物。对于稀释样品，保持Russell毒蛇毒因子X激活剂抗体和捕获抗体的浓度恒定，并在不使标记抗体浓度恒定的情况下将复合物稀释到缓冲液中，等同于检测低神经毒素浓度。此ELISA-酶联凝血法是小鼠生物测定法的一种方便替代方法，除了必要的敏感性外，该方法还显示了神经毒素的完全分解。

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