Detection and quantitation of chloramphenicol by competitive enzyme-linked immunoassay

A competitive enzyme-linked immunoassay for the detection and quantitation of chloramphenicol has been developed. The binding of specific rabbit antibody to solid-phase-bound chloramphenicol was competitively inhibited by free chloramphenicol in the sample to be assayed. Antibody not displaced was indicated by using a commercially available, enzyme-linked, anti-rabbit immunoglobulin preparation and reacted with added substrate. Enzyme activity, measured spectrophotometrically, was inversely proportional to the concentration of chloramphenicol in the sample. Quantitation of the antibiotic was linear to 100 ng/ml, with a lower limit of detection of 1 ng/ml (P less than 0.05). Specificity was demonstrated by the lack of inhibition by any of 31 selected antimicrobial agents or chemicals tested in the assay. Chloramphenicol sodium succinate and thiamphenicol, an experimental antibiotic similar in structure to chloramphenicol, were the only drugs found to produce cross-reactions. In addition to excellent sensitivity and specificity, the assay was shown to have good precision and economy and could be completed in approximately 24h.