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The ovarian cholesterol depletion assay for luteinizing hormone (LH) was investigated in the untreated immature rat. The prepubertal rat is a satisfactory animal to use in this assay because the diurnal rhythm in ovarian cholesterol is not present until puberty. A dose-response curve was obtained between 2·5 and 10·0 µ LH and the average lambda was 0·17. The assay was specific for LH; the presence of other pituitary hormones had no significant effect on the relative potency of NIH-LH preparations. The assay is probably not sensitive enough for untreated plasma but had excellent reproducibility and is easily performed.