RESUMO
AIMS: To evaluate the sensitivity and specificity of polyclonal and monoclonal antibodies (Mabs) against intimin in the detection of enteropathogenic and enterohaemorrhagic Escherichia coli isolates using immunoblotting. METHODS AND RESULTS: Polyclonal and Mabs against the intimin-conserved region were raised, and their reactivities were compared in enteropathogenic E. coli (EPEC) and enterohaemorrhagic E. coli (EHEC) isolates using immunoblotting analysis. In comparison with rat antiserum, rabbit anti-intimin IgG-enriched fraction had a stronger recognition pattern to a wide spectrum of intimin types in different EPEC and EHEC serotypes. On the other hand, murine monoclonal IgG2b specific to intimin, with dissociation constant of 1.3x10(-8) mol l(-1), failed in the detection of some of these isolates. CONCLUSION: All employed antibodies showed 100% specificity, not reacting with any of the eae-negative isolates. The sensitivity range was according to the employed antisera, and 97% for rabbit anti-intimin IgG-enriched fraction, followed by 92% and 78% sensitivity with rat antisera and Mab. SIGNIFICANCE AND IMPACT OF THE STUDY: The rabbit anti-intimin IgG-enriched fraction in immunoblotting analysis is a useful tool for EPEC and EHEC diagnoses.
Assuntos
Adesinas Bacterianas/imunologia , Anticorpos Monoclonais/imunologia , Escherichia coli Êntero-Hemorrágica/classificação , Escherichia coli Enteropatogênica/classificação , Proteínas de Escherichia coli/imunologia , Animais , Especificidade de Anticorpos , Feminino , Soros Imunes/imunologia , Immunoblotting/métodos , Imunoglobulina G/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Coelhos , Ratos , Sensibilidade e EspecificidadeRESUMO
To evaluate the sensitivity and specificity of polyclonal and monoclonalantibodies (Mabs) against intimin in the detection of enteropathogenic andenterohaemorrhagic Escherichia coli isolates using immunoblotting.Polyclonal and Mabs against the intimin-conservedregion were raised, and their reactivities were compared in enteropathogenic E. coli (EPEC) and enterohaemorrhagic E. coli (EHEC) isolates using immunoblotting analysis. In comparison with rat antiserum, rabbit anti-intimin IgG-enriched fraction had a stronger recognition pattern to a wide spectrum of intimin types in different EPEC and EHEC serotypes. On the other hand, murine monoclonal IgG2b specific to intimin, with dissociation constant of1Æ3 · 10)8 mol l)1, failed in the detection of some of these isolates. All employed antibodies showed 100% specificity, not reacting with any of the eae-negative isolates. The sensitivity range was according to the employed antisera, and 97% for rabbit anti-intimin IgG-enriched fraction, followed by 92% and 78% sensitivity with rat antisera and Mab. Significance and Impact of the Study: The rabbit anti-intimin IgG-enriched fraction in immunoblotting analysis is a useful tool for EPEC and EHEC diagnoses.