Radioimmunoassay of rat luteinizing hormone (LH) in the sera and the anterior pituitaries of normal and castrated males and proestrous females employing different batches of anti-LH sera gave the results suggesting heterogeneity of immunoreactive LH (IR-LH). Gel filtration analyses of anterior pituitary extract and pooled sera failed to give positive evidence for the heterogeneity. But when the extracts of normal rat anterior pituitaries were electrofocused and the fractions were assayed, the pituitary glycoprotein hormones, LH, FSH and TSH, were found to consist of several components with different isoelectric points. In electrofocusing patterns of IR-LH, distinct peaks were observed at pH 8.5 (component A), 8.8 (component B), 9.1 (component C) and 9.3-9.4 (component D), and less distinct peaks were observed at pH 7.9 (component A'), 9.6 (component E) and 9.8 (component F). There seemed to be some sexual difference in the relative amounts of these components. Orchiectomy caused marked changes in electrofocusing patterns of IR-LH, i.e., increase of components A', A, B, C and D. The blockade of LH release by estradiol benzoate and progesterone treatment in orchiectomized rats caused a marked increase of component A. At the proestrous LH surge, IR-LH components were evenly released. Radioimmunoassay of IR-LH components with multiple antisera indicated that these antisera had different affinities to these components.