C57BL/6 mice deficient in D^b or K^b and were generated by gene targeting in the laboratory of Dr. Francois Lemonnier of The Institut Pasteur, Paris France. Generation of H2-D^b knockout mice: The H2-D^b gene was disrupted using an insert containing LacZ and a neo cassette flanked by a 4-kb HindIII-XbaI 5' and 1-kb KpnI-SpeI 3' fragment of the D gene resulting in the loss of the first three exons. The targeting construct was introduced into a CGR-8 embryonic stem cell of 129/OLA origin. Clones that were confirmed for homologous recombination were injected into B6 blastocysts and germline chimeras were identified. The founders were crossed twice with C57BL/6Ji females and heterozygous N2 offspring were intercrossed to generate the H2-D^b homozygous strain. Inactivation of the H2-Db gene was confirmed by Southern blot analysis on tail DNA and immunofluorescence assay. Generation of H2-D^b knockout mice: the H2-D^b gene was disrupted by replacing the 2nd and 3rd exons with an HPRT minigene inserted in the opposite transcriptional orientation.The disrupted gene was introduced into an E14TGa(H-2b and b2m) HPRT deficient embryonic stem cell of 129/OLA origin. After injection into B6 blastocysts, the resultant germline chimeras were backcrossed twice to C57BL/6Ji mice and offspring heterozygous for the H2-K^b knockout allele were crossed to generate H2-K^b deficient mice (K^b -/-). Inactivation of the H2-K^b gene was confirmed by Southern blot analysis on tail DNA and by FACS analysis. Generation of H2-K^b H2-D^b double knockout mice: N2 K^b -/- and D^b -/ - mice were intercrossed and the H2-K^b and H2-D^b targeted mutations were genetically linked by crossing-over between the two mutated loci. The double targeted mutation mice were subsequently backcrossed to C57BL/6Ji mice another 10 times