Physiological levels of luteinizing hormone (LH), in concert with follicle stimulating hormone (FSH), promote ovarian follicular development and ovulation. However, high LH levels associated with ovarian dysfunction have been shown to inhibit these processes. Thus, developing a selective LH inhibitor could be potentially useful for treating ovarian dysfunction. Here, we developed a mouse LH-binding protein (mLBP) composed of the extracellular domain of LH receptors as a selective inhibitor of mouse LH. After transient introduction of mLBP expressing vectors into Expi293F cells, mLBP was obtained as a soluble protein via a cleavage reaction with thrombin. The binding ability of mLBP for mouse LH was confirmed using sera containing high LH and FSH collected from ovariectomized (OVX) mice. The bioactivity of mLBP was demonstrated by inhibition of cAMP and testosterone productions induced by OVXmouse serum in mouse Leydig MLTC-1 cells expressing LH receptors. In contrast, mLBP did not bind mouse FSH and inhibit cAMP production induced by OVX-mouse serum in 293 cells expressing mouse FSH receptors. The mLBP also showed binding affinity to human LH (hLH), and inhibited hLH-induced cAMP production in MLTC-1 cells. Thus, the mLBP selectively suppresses the action of LH and is a potential therapeutic agent for ovarian dysfunction.