Homogenates of human pancreas in saline were centrifuged at 27 000 X g and the supernates were fractionated by preparative polyacrylamide gel electrophoresis. The gels were divided into sections and each section was injected into rabbits; after absorption with polymerized serum from apparently normal humans, the antiserum obtained by injecting one of the sections was tested against a variety of human tissue extracts but reacted only with saline extracts of human pancreas. The absorbed antiserum, polymerized and made insoluble with glutaraldehyde, was used to purify a pancreas-specific antigen by immunoaffinity batch technique. The purified antigen proved to be a protein with some carbohydrate content (180 mg/g by weight) and a molecular mass of about 2.25 X 10(5) daltons. The antigen is relatively thermostable, and precipitates in the range of 245.64-340.2 g/L saturated ammonium sulfate; its antigenic activity is not affected by incubation with ribonuclease or deoxyribonuclease, but is destroyed by incubation with trypsin or neuraminidase and by extraction with perchloric acid. Immunofluorescence studies show that the antigen is diffusely present in the cytoplasm of pancreatic acinar cells.