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首页> 外文期刊>Journal of Clinical Microbiology >Stool desorbing activity: a possible cause of false-positive reactions in competitive enzyme immunoassays.
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Stool desorbing activity: a possible cause of false-positive reactions in competitive enzyme immunoassays.

机译：粪便解吸活性：竞争性酶免疫法中假阳性反应的可能原因。

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We have developed a competitive enzyme immunoassay for the measurement of purified toxin A of Clostridium difficile. However, when we applied this assay to the detection of C. difficile toxin in stool specimens, we noted a high rate of nonspecific activity in fecal specimens which did not contain toxin. We found that the low specificity (26%) of the assay was due to the presence in stool specimens of interfering factors which desorbed the antigen coated on the solid-phase surface. These factors could be detected by measurement of the desorption of biotin-labeled proteins attached to the solid-phase surface. In addition, these interfering factors were partially inactivated by heating at 56 degrees C for 10 min and partially inhibited by phenylmethylsulfonyl fluoride (2 mM) or soybean trypsin inhibitor (10 mg/ml). These data suggested that the desorbing activity was due to proteolytic activity in the fecal specimens. Fetal calf serum (50%) was found to be the most effective measure in preventing the interfering effect. By using 50% fetal calf serum as a diluent, we increased the specificity of the antibody inhibition enzyme immunoassay to 93%. Interfering factors in stool specimens could be a cause of false-positive results in other competitive immunoassay systems. The use of diluents which neutralize protease activity can result in a marked improvement in the specificity of competitive immunoassay systems.
机译：我们已经开发了一种竞争性酶免疫法，用于测定艰难梭菌纯化毒素A。但是，当我们将此测定法应用于粪便标本中艰难梭菌毒素的检测时，我们注意到粪便标本中不含毒素的非特异性活性很高。我们发现该测定法的低特异性（26％）是由于粪便标本中存在干扰因子，该干扰因子解吸了固相表面上包被的抗原。这些因子可以通过测量附着在固相表面的生物素标记蛋白的解吸来检测。此外，这些干扰因子在56°C加热10分钟后部分失活，并被苯甲基磺酰氟（2 mM）或大豆胰蛋白酶抑制剂（10 mg / ml）部分抑制。这些数据表明解吸活性是由于粪便样品中的蛋白水解活性所致。发现胎牛血清（50％）是预防干扰作用的最有效措施。通过使用50％的胎牛血清作为稀释剂，我们将抗体抑制酶免疫测定的特异性提高到93％。粪便标本中的干扰因素可能是其他竞争性免疫分析系统中假阳性结果的原因。使用中和蛋白酶活性的稀释剂可导致竞争性免疫分析系统的特异性显着提高。

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