Development of a radioimmunoassay for Escherichia coli heat-stable enterotoxin: comparison with the suckling mouse bioassay

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Abstract
E. coli strains which produce heat-stable enterotoxin (ST) are usually identified by demonstrating the production of ST. At present, ST can be detected only by bioassay methods. Recently, E. coli ST was purified, enabling development of a radioimmunoassay for ST. Radioiodination of ST was performed by the lactoperoxidase method, which resulted in a high specific activity and retained the biological activity of ST. Anti-ST antisera were raised in goats by injecting the goats with pure ST coupled to bovine IgG. Antibody titers ranged from 1:8000 to 1:40,000. Using these reagents, assay conditions were examined thoroughly. A 14- to 18-h incubation at 4.degree. C in sodium acetate buffer with an ionic strength of 120 mM (pH 6.2) gave maximal sensitivity and reproducibility. Free ST was separated from antibody-bound ST by dextran-coated charcoal. This radioimmunoassay accurately and reproducibly measured ST in the range from 50-500 pg ST/tube and could quantitate ST accurately in complex bacteriological media. This assay was specific for STa, measured human and porcine STa equally well and did not cross-react with STb, with several other enterotoxins or with various gastrointestinal peptides. Intact disulfide bridges in the ST molecule were required for immunoreactive activity.