Stool desorbing activity: A possible cause of false-positive reactions in competitive enzyme immunoassays

M. Hanvanich, R. Viscidi, B. E. Laughon, J. G. Bartlett, R. H. Yolken

Research output: Contribution to journalArticle

Abstract

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° 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.

Original languageEnglish (US)
Pages (from-to)184-188
Number of pages5
JournalJournal of Clinical Microbiology
Volume21
Issue number2
Publication statusPublished - 1985

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ASJC Scopus subject areas

  • Microbiology (medical)
  • Microbiology

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