Sensitive microplate assay for detection of bactericidal antibodies to Vibrio cholerae O139

Abstract

A microplate assay for the detection of bactericidal antibodies to Vibrio cholerae O139 is described. The assay is sensitive, highly reproducible, specific, and convenient to perform. It has been used to demonstrate the induction of serum bactericidal antibodies in Vietnamese recipients of an oral, inactivated, bivalent O1/O139 vaccine, as well as in Bangladeshi patients with O139 disease. In both study groups there was a significant inverse correlation between the preexposure level of antibodies in serum and the magnitude of the subsequent bactericidal response. Although infection generated stronger responses than vaccination, the proportion of responders was similar among individuals with low background titers.

FIG. 1.

Reproducibility of modified microplate assay protocol. (A) Eight sera were assayed on five separate occasions; the titers of six showed no variability (e.g., samples A and C), while the other two (samples B and D) showed a twofold range in titer. (B) Three serum pairs (X to Z) were each titrated three to four times, with the results shown. The titer preexposure is shown in black at the bottom of each histogram; the postexposure titer is represented by the open histogram. The ratio of postexposure to preexposure titers remained constant.

FIG. 2.

Magnitude of anti-O139 serum bactericidal responses induced by vaccination or disease in relation to preexisting antibody titers. (A) Responses (fold rise in titer) of Vietnamese recipients of inactivated O139 vaccine, plotted in relation to prevaccination antibody titers. (B) Responses (titer of the convalescent serum divided by the titer of the acute-phase serum) of Bangladeshi cholera patients, plotted in relation to acute-phase antibody titers.