DNA was isolated from 75 mouse strains carrying classical H-2 haplotypes as well as haplotypes derived from wild mice. The DNA was digested with three restriction endonucleases, Bst EII, Eco RI, and Bam HI, the digests hybridized, using the Southern blotting technique, with probes for the class II genes As, As, E~, and E~, and the restriction fragment length polymorphism at these loci determined. The analysis revealed that the most polymorphic of the four loci is As, followed by E~, and, at a different level, by E~ and A s. There is a large difference in the degree of polymorphism between the A s and Ep genes, on the one hand, and the As and E~ genes, on the other hand. There is no difference in the degree of polymorphism between the A s and E~ genes. These findings do not substantiate previous postulates of a high A s polymorphism and they do not agree with the hypothesis that the class II region is divided into highly polymorphic centromeric and less polymorphic telomeric subregions. Rather, it appears that the differences in the degree of polymorphism of the different segments of the class II region are determined by the class II loci themselves. The polymorphism of the less polymorphic class II genes is, however, still greater than the polymorphism of certain other genes on chromosome 17, notably the c~4-globin pseudogene. The distribution of polymorphisms at the A s and E~ loci suggests that even populations occupying relatively small geographical regions differ in alleles at these loci. Sharing of A s alleles between unrelated populations is yet to be detected. A certain degree of linkage disequilibrium exists among the A,, As, and E~ loci; by contrast, the E, locus appears to vary largely independently of the other class II loci.