Detection of Cholera Toxin by a Highly Sensitive Bead‐Enzyme Linked Immunosorbent Assay

Abstract

A bead-enzyme linked immunosorbent assay (bead-ELISA) for detection and quantification of cholera toxin (CT) in broth cultures of Vibrio cholerae O1 has been developed. Under optimal buffer and pH conditions the bead-ELISA could consistently detect 40 pg/ml of CT. None of the ingredients of commonly used media for in vitro culture of V. cholerae O1 hindered the performance of the bead-ELISA. Evaluation of the sensitivity and specificity of the bead-ELISA against the commonly used reversed passive latex agglutination (RPLA) test for detection of CT was performed using a collection of 239 strains of V. cholerae O1 (including both biotypes and serotypes) which were examined by a gene probe encoding for the A1 subunit of CT. Although both the assays were highly specific, the bead-ELISA was more sensitive than the RPLA. Quantification of CT by the bead-ELISA revealed that the concentration of CT produced by the strains of V.which were negative by the RPLA was lower than 1 ng/ml and therefore below the minimum detection ability of the RPLA. The bead-ELISA is a simple, specific and highly sensitive assay for routine detection of CT and is recommended for routine use in clinical microbiology laboratories.

Incidence and toxigenicity of Vibrio cholerae in a freshwater lake during the epidemic of cholera caused by serogroup O139 Bengal in Calcutta, India

“…Confirmed strains of V. cholerae O139 and all strains of V. cholerae that did not agglutinate with O1 or O139 antisera were examined for production of cholera toxin (CT) by a highly sensitive bead enzyme-linked immunosorbent assay [21]. The strains were grown in casamino acid yeast extract medium supplemented with 90 #g ml -t of lincomycin (Sigma Chemical Co., St. Louis, MO) using stationary culture at 30°C in 90 x 16 mm Petri dishes [22].…”

Development and evaluation of a multiplex PCR assay for rapid detection of toxigenicVibrio choleraeO1 and O139

“…Realizing the need for quick diagnosis, a variety of rapid tests including co-agglutination tests [11^13], enzyme-linked immunosorbent assays [14,15] and membrane based immunoassays [16] have been developed in the recent past. Recently, Albert et al [17] have developed a polymerase chain reaction (PCR) assay using a primer pair corresponding to a chromosomal region of V. cholerae O139 for rapid detection of the serogroup from stool samples.…”

Typing of Verotoxins by DNA Colony Hybridization with Poly‐ and Oligonucleotide Probes, a Bead‐Enzyme‐Linked Immunosorbent Assay, and Polymerase Chain Reaction

“…VTEC strains used in this study were isolated from either humans or animals and are listed in Table 1. The standard E. coli strains carrying cloned VT1 (11), VT2 (31), VT2vha (7), and VT2vhb genes (7 The bead-ELISAs developed for VT1 and VT2 were as sensitive as that for the cholera toxin reported previously (19,30). As little as 60 pgiml of the toxin was detected (Fig.…”