High titer antibodies to secretin were produced in rabbits by immunization with purified porcine secretin (from GIH) conjugated to bovine serum albumin in complete Freund's adjuvant. Synthetic porcine secretin (from Squibb) was radiolabeled with 125I using chloramine-T and radiosecretin was purified using talc-silica and fibrocellulose powder. Specificity of antibodies to secretin was extensively evaluated in studies examining (1) cross-reactivity with other naturally occurring hormones, (2) parellelism of serum dilutions and secretin standards, (3) quantitative recovery of secretin added to serum samples, and (4) changes in circulating immunoreactive secretin in response to intraduodenal acidification. The mean fasting serum secretin concentration in 10 dogs was 65 +/- 5 (SEM) pg per ml and in 21 healthy human volunteers it was 69 +/- 5 pg per ml. In response to intraduodenal instillation of 10 mEq of HCl, portal venous secretin concentrations increased from a mean fasting value of 52 to 450 pg per ml within 2 min; peripheral venous secretin increased from 66 to 229 pg per ml within 5 min. No significant changes in peripheral serum immunoreactive secretin concentrations were identified in dogs after a protein meal. Intravenously administered secretin in man disappeared from the circulation with mean half-life 4.06 +/- 0.82 (SEM) min.