Efficient transport of class I major histocompatibility complex molecules to the cell surface requires association of the class I heavy chain with endogenous peptide and the class I light chain, beta 2-microglobulin (beta 2M). A mutant cell line deficient in beta 2M transports low amounts of nonpeptide-associated heavy chains to the cell surface that can associate with exogenously provided beta 2M and synthetic peptide antigens. Normal beta 2M-sufficient cells grown in serum-free media devoid of beta 2M also require an exogenous source of beta 2M to efficiently bind synthetic peptide. Thus, class I molecules on normal cells do not spontaneously bind or exchange peptides.