Aldose reductase (AR) was purified from rat and bovine seminal vesicles using DEAE-cellulose, hydroxylapatite, and Sephadex-gel column chromatography. The purification resulted in the obtention of an AR pool and a contaminating pool. Antibodies were raised in rabbits against both enzymes by subcutaneous injection of the AR pool. The antisera was judged to be specific for AR by immunoprecipitation of AR activity and by Ouchterlony double immunodiffusion and immunoelectrophoretic methods. Antibodies against rat AR were used in the unlabeled antibody-enzyme (PAP) technique to demonstrate the cellular location of the enzyme in a number of tissues known to be sites of diabetic lesions. Antibodies against bovine AR were not cross reactive with the rat enzyme, as determined by Ouchterlony and competitive protein-binding studies. AR was localized in rat tissues to the Schwann cell sheath of peripheral nerve, arterial endothelium, and the sustentacular (Sertoli) cells and mature spermatids of the testis.