F1 hybrid mice often reject parental hematopoietic grafts, a phenomenon known as hybrid resistance. Hybrid resistance is mediated by natural killer (NK) cells and although the molecular interactions responsible for this phenomenon are largely unknown, one hypothesis suggests that parental cells are rejected because they fail to express a complete set of host major histocompatibility complex (MHC) class I molecules. Inherent in this theory is that NK cells in the F1 hybrid are instructed by self MHC class I molecules to form an NK cell repertoire capable of reacting against cells lacking these self MHC class I molecules. Here, we show that C57BL/6 x DBA/2 mice (H-2b/d) devoid of beta2-microglobulin (beta2m) are incapable of rejecting beta2m-/- parental C57BL/6 cells (H-2b) both in vivo and in vitro. From this, we conclude that the development of an NK cell repertoire, at least in F1 mice of the H-2b/d haplotype, requires expression of MHC class I molecules complexed with beta2m.