The maximum genetic diversity of horsegram (Macrotyloma uniflorum) is considered to be in the Old World tropics, especially in India and the Himalayas. Identification of horsegram genotypes is of prime importance to ensure quality seed by identifying off-types in samples. Characterization based on morphological characters usually varies with environments, and evaluation of these traits requires growing the plant to maturity prior to identification. Therefore, 22 genotypes of horsegram were identified based on seed protein profile and isozyme polymorphism. Most genotypes were identified successfully based on genotype specific intensity of dark, medium and light bands as well as their relative position in the seed protein and isozyme profiles analyzed by the polyacrylamide gel electrophoresis. Their characterizations were difficult through the total number of bands and as several of them were common in more than one genotype. Among the isozymes studied esterase isozyme profile is found to be more accurate for genotype characterization.