Simple and rapid purification procedures for parvalbumin, one of the Ca2+-binding proteins (extracted from rat skeletal muscle), were developed, and its antiserum was produced in rabbits to measure the parvalbumin content of various rat tissues by RIA. The heat treatment, ammonium sulfate fractionation, and trichloroacetic acid precipitation of soluble fraction from rat skeletal muscle followed by single diethylaminoethyl Sephadex A-50 column chromatography yielded a pure 120 mg protein from 150 g skeletal muscle. Amino acid analysis, together with electrophoretic mobility, indicated that the protein was identical to parvalbumin. The antisera to this rat skeletal muscle parvalbumin (raised in rabbits) did not cross-react with calmodulin or S-100 proteins. The RIA for parvalbumin using this antisera and [125I]parvalbumin revealed that skeletal muscle and brain contained high levels of the antigen; the values of which were 69,486 +/- 4,933.1 and 881 +/- 165.6 ng/mg protein, respectively. However, the parvalbumin antigen in the heart, lung, liver, and spleen could not be detected. On the other hand, the contents of the antigen in the endocrine glands (in nanograms per mg protein) were as follows: pituitary (125 +/- 46.6), thyroid (108 +/- 50.0), adrenal (341 +/- 64.3), testes (227 +/- 37.2), and ovaries (218 +/- 10.1). All of these values were comparable to levels of antigen found in the brain sample. These results suggest an important role for parvalbumin in endocrine glands.