A rat testes tubule receptor preparation has been characterized with respect to its usefulness for measurement of FSH activity in fractions derived from human pituitary glands. Highly purified human FSH, iodinated by a modification of the chloramine-T procedure, was employed as the radioligand. A series of human gonadotropin fractions were assayed in the FSH radioligand-receptor assay (RLA) and also in the Steelman-Pohley hCG-augmented rat ovarian weight gain assay (S-P assay), using LER-907 (20 IU/mg) as the assay reference preparation in each instance. The index of discrimination (ID) for a series of hFSH fractions ranging in potency from 53 IU/mg to 3608 IU/mg was 1.05. Highly purified hLH, hTSH and hCG showed 125I-hFSH uptake inhibition activity ranging from 0.9 to 3.6% that of highly purified hFSH. Significant uptake inhibition of labeled hFSH could be obtained with 2.5 ng of pure hFSH, while no uptake inhibition was seen with 10,000 ng of human growth hormone, human prolactin, ovine prolactin, ACTH and human serum albumin. Asialo-hFSH had a potency in the RLA 680 times greater than in the S-P assay, indicating that as with hLH and hCG, sialic acid is not required for receptor binding activity of hFSH. The usefulness of the RLA in following the combination of subunits of hFSH was also demonstrated. (Endocrinology94: 483, 1974)