Abstract Twelve mouse monoclonal antibodies for human luteinising hormone were produced. The affinities varied from 4 × 10 7 to 1 × 10 10 1/mol. The specificity of each antibody was assessed by determining the relative reactivities with luteinising hormone, thyroid stimulating hormone, follicle stimulating hormone and chorionic gonadotrophin. Six antibodies bound to the α-subunit as shown by similar reactivity with all hormones, and the remainder to the β-subunit as shown by specificity for luteinising hormone. This latter group of antibodies cross-reacted only weakly with thyroid stimulating hormone (approximately 10%) and follicle stimulating hormone (approximately 3%). Three of these antibodies also showed low reactivity towards chorionic gonadotrophin (< 10%), though the others did not (80–300%). The ability of different antibodies to bind simultaneously to luteinising hormone was examined and it was shown that several distinct antigenic determinants existed on both subunits. The characterisation of monoclonal antibody binding sites is discussed in relation to the use of antibodies in two-site immunoradiometric assays.