Four groups of four healthy mid-lactation Friesian cows were fed a compound feedingstuff containing either 2, 10 or 250 mg sulphamethazine/kg, corresponding to 0, 2, 10 and 250 per cent of the therapeutic inclusion rate in rations for pigs, at a flat rate of 3 kg twice daily for 21 days, followed by a seven-day withdrawal period. The cows were machine-milked twice daily and pooled milk samples from each cow were analysed by a commercially available microbiological assay with a sensitivity of 100 μg/litre and by a high performance liquid chromatography (HPLC) procedure with a limit of detection of 10 μg/litre. No sulphamethazine was detected by HPLC in the milk samples taken from any of the cows fed the concentrate containing 2 or 10 mg/kg. The milk samples from all four cows fed the highest concentration of sulphamethazine contained from 21 to 120 μg/litre while they were being fed the contaminated concentrate. The cow with the highest concentrations of sulphamethazine was the only one which repeatedly tested positive by the microbiological assay. The concentration of sulphamethazine declined rapidly during the withdrawal period and the drug was not detectable by either method in samples taken from two days after the contaminated feed was withdrawn.