The concentration of the new bioactive acetic seminal fluid protein (aSFP) was determined in body fluids of bulls. An aSFP-specific radioimmunoassay was elaborated and established for routine measurement. No crossreaction of the aSFP antibody was found with other seminal plasma proteins and known growth factors. The 14 kDa aSFP was detected in secretions of ampulla and seminal vesicles by immunoblot analysis, but not in testis, epididymis or blood. Large quantities of aSFP were measured by radioimmunoassay in the fluid from the ampulla (2.6 +/- 0.3 mg ml-1) and seminal vesicles (3.0 +/- 0.4 mg ml-1). Immunohistochemistry techniques demonstrated that aSFP was localized mainly in the secretory epithelium of ampulla and seminal vesicles. Large amounts of aSFP (4.0 +/- 0.4 mg ml-1) were present in seminal. plasma of bulls within a range of 1-7 mg ml-1, but aSFP could not be found on membranes of spermatozoa. Concentrations vary considerably among individuals, suggesting that aSFP may have a role in bovine reproduction.