A radioimmunoassay procedure for the determination of pirenzepine in either plasma or urine was demonstrated to be both sensitive and specific as well as highly reproducible. The assay could detect levels as low as 1.25 ng/ml. The sensitivity was sufficient to allow the analysis of biological samples from both pharmacokinetic and clinical studies. The two metabolites of pirenzepine, LS 75 and LS 822, did not cross-react with the antiserum. The assay was not affected by a change in anticoagulant or by the presence of several over-the-counter or prescription drugs, even at very high levels. Samples could be frozen and stored for at least a year without affecting the analysis. Repeat analysis could be performed on samples that had been refrozen. Several thousand plasma and urine samples, including plasma samples from severely renally impaired patients, have been analyzed for pirenzepine by the RIA with no interferences having been detected.