Determination of clenbuterol in bovine tissues and urine by enzyme immunoassay

We have developed an enzyme immunoassay (EIA) for clenbuterol using an antiserum raised in rabbits by immunization against a clenbuterol diazo derivative coupled to human serum albumin. Horseradish peroxidase coupled to the clenbuterol diazo derivative was selected as a tracer. The dose of clenbuterol that caused 50% binding inhibition was 18 pg per well, and the limits of detection of the EIA were 0.15 and 0.3 ppb in urine and liver, respectively (when the variability of blank values in samples from untreated animals was taken into account). This assay was used to test urine and tissue samples collected from veal calves and cows that had received in their feed a number of concentrations of clenbuterol over a range of time periods