Polyclonal antibodies against the coccidiostat, lasalocid, were raised in sheep. The antisera were applied in an enzyme‐linked immunosorbent assay (ELISA) for lasalocid, which was validated for chicken serum, liver and muscle. Bridge homology in the ELISA was overcome by absorbing unspeciflc antisera onto a conjugate between salinomycin and chicken serum albumin, which was immobilized onto Biosilon beads. The described assay is highly specific for lasalocid, and is capable of detecting lasalocid at concentrations of less than 015 ng/g, depending on the tissue. Residual concentrations of lasalocid in broilers fed medicated feed containing 90 mg/kg lasalocid were measured during a 7 day withdrawal period. The half‐life of lasalocid was 11, 36 and 41 h in serum, liver and muscle, respectively. Lasalocid concentrations in liver were approximately 10 ng/g 7 days withdrawal of the medicated feed.