The relationship among different minor lymphocyte stimulatory locus (Mls) determinants, Mlsa, Mlsb, Mlsc and Mlsd, remains unclear. Because of the high degree of cross-reactivity between Mlsa and Mlsd determinants, the weak stimulatory capacity of Mlsc, and the generally non-stimulatory nature of Mlsb, some investigators have recently suggested that Mls is composed of only a single expressed allele originally defined as the a and d alleles. In order to clarify the nature of Mls determinants, T cell clones positively selected for reactivity to the three stimulatory Mls determinants, Mlsa, Mlsc and Mlsd, were generated and their specificities defined by extensive genetic studies. The response pattern of these Mls-specific clones as well as the results of primary mixed lymphocyte responses indicated that: (i) Mlsa and Mlsc determinants recognized by T cells are distinct, demonstrating that polymorphism does exist within the Mls system; and (ii) Mlsd is not an independent Mls type since Mlsd determinants are composed of Mlsa and Mlsc determinants and anti-Mlsd T cell responses are, in fact, the sum of anti-Mlsa and anti-Mlsc responses.