The properties of a synthetically produced peptide composed of the same primary structure of 18 amino acids described for human Escherichia coli heat-stable enterotoxin were compared with those of purified heat-stable toxin obtained by bacterial growth. The dosage required to evoke fluid secretion in the suckling mouse and rat ligated ileal loop assays was the same for both toxins. The antigenicity of the two toxins was similar when assayed by enzyme-linked immunosorbent assay with hyperimmune antiserum to either toxin. The secretory effect of the two toxins in the suckling mouse assay was seroneutralized by the same dilutions of hyperimmune antiserum to either toxin. Immunization of rats with the synthetic toxin coupled to a large-molecular-weight carrier raised serum and mucosal antitoxin responses which provided protection against challenge with either the synthetic or biological toxin as well as with viable heat-stable enterotoxin-in-producing organisms. These observations indicate that synthetically produced heat-stable toxin has the same properties as the toxin derived by bacterial culture. The availability of the more readily made synthetic form of heat-stable toxin should facilitate the production of a vaccine based on cross-linking this toxin with either the heat-labile toxin or its nontoxic B subunit.