Little is known about the nature of recombination hotspots in the human genome and the relationship between crossover activity and patterns of linkage disequilibrium. We have therefore used both haplotype analysis and direct detection of crossovers in sperm to characterize a putative recombination hotspot in the TAP2 gene within the class II region of the MHC. Haplotype diversity provided evidence for a localized hotspot within intron 2 of this gene. Sperm DNA typing using allele-specific PCR primers to selectively amplify recombinant TAP2 molecules revealed a highly localized meiotic crossover hotspot approximately 1.2 kb long, unusually abundant in sequence polymorphisms and flanked by DNA much less active in recombination. Sperm crossover appeared to be fully reciprocal, and almost all crossover products were simple, involving a single exchange between adjacent heterozygous markers. This hotspot appears to be much more active in female than male meiosis. No primary sequence similarities could be found between any of the very few well defined crossover hotspots in the human genome, all of which show recombinationally active domains 1-2 kb long. Direct comparison of recombination frequency and haplotype diversity in TAP2 showed that linkage disequilibrium measures were a poor predictor of crossover frequency in this region, with non-recombining markers sometimes in free association and with examples of pairs of markers spanning the recombination hotspot showing substantial or even absolute linkage disequilibrium.