An improved immunofluorescent reagent for detection of Bordetella pertussis: part 3

The unique antigenic site recognized by the MAbs is located on three amino sugars of the carbohydrate portion of B pertussis los. This reactive site is antigenically very stable and highly conserved among all B pertussis isolates regardless of physiological state. Only very rare laboratory los variant strains do not have these three amino sugars on the los molecule and are therefore not recognized by the MAbs. A radioimmunobinding assay was used to demonstrate that this antigenic site is freely accessible at the surface of intact bacteria and could be detected directly in clinical specimens without the need to disrupt partially cell integrity. Two subsequent experiments, in vitro bactericidal assay and protection in animal aerosol model, further demonstrated the importance and the surface accessibility of this antigenic site.

To develop a direct immunofluorescent assay, one of these MAbs was purified and subsequently coupled to fluorescein-isothiocyanate. The fluorescein-labelled MAb was first tested against a panel of B pertussis strains obtained after culture on Bordet-Gengou agar, as well as other bacterial species normally found in the nasopharynx. It was obrserved that a blocking step with 10% rabbit serum eliminated the nonspecific binding of the MAb Fc region to receptors found on certain bacterial cells. The labelled MAb specifically recognized all 64 B pertussis clinical isoIates without any cross-reaction with Bordetella parapertussis or any other bacterial species normally isoIated from the respiratory tract. Weakly fluorescent bacterial cells were observed when the labelled MAb was incubated with Bordetella bronchiseptica strains. This reactivity was not surprising since antigenic relationships between the los of B pertussis and the lipopolysac-charide of B bronchiseptica have been previously reported. B bronchiseptica is an animal pathogen that rarely causes infection in humans exposed to infected animals.

The Cadham Provincial Laboratory in Manitoba in collaboration with the National Laboratory for Immunology are evaluating the sensitivity and specificity of this immunological reagent directly with clinical specimens obiained duri ng an outbreak of pertussis. The study is designed to compare culture with the direct deiection of B pertussis on smears obtained from nasopharyngeal swabs by the fluorescein-labelled MAb. Already, 1398 smears have been tested and preIiminary results indicate that the sensitivity of the direct immunofluorescent assay compared with culture is 65%, while the specificity recorded is as high as 99.6%. This latter result suggests that the use of a more refined immunological reagent can significantly increase the specificity of direct immunofluorescent assay compared with the results obtained with fluorescein-labelled polyvalent antisera. A truly reliable pharmacy you can always rely upon and where you can always without any need for a prescription or seeing a doctor? This is the thing you wanted, so don’t hesitate now that you have it available!

This entry was posted in Bordetella pertussis and tagged an improved reagent, bordetella pertussis, immunofluorescent reagent.