Graves' disease (GD) is caused by genetic and environmental factors. To date only the HLA class II region on chromosome 6p21 and the CTLA-4 gene on chromosome 2q33 have been consistently associated with disease. A recent study has indicated the most likely position of the aetiological variant in the CTLA-4 locus to be in a 6.1kb region of the 3' untranslated region of the gene. However the same degree of mapping resolution has not been achieved for the HLA class II region due to the strong linkage disequilibrium between the DRB1, DQB1 and DQA1 loci. The aim of this study was to fine-map the primary locus involved in the development of GD within the HLA class II region. Eight hundred and seventy one patients with GD and 621 control subjects were genotyped at the DRB1, DQB1 and DQA1 loci using the polymerase chain reaction. Local ethics approval was obtained for this study and all patients gave informed, written consent. All three loci were associated with GD, P = 1.45 x 10-12, 3.2 x 10-5 & 9.26 x 10-12, respectively. Stepwise logistic regression analysis showed that the association could be explained by either DRB1 or DQA1. Since previous studies have implicated position beta74 of DRB1 in susceptibility to GD, we predicted the amino acid sequence of the exon two-encoded peptide binding domain of DRB1 and analysed each position for association with GD. Thirteen amino acids were associated with GD (P = 2.2 x 10-4- 1.2 x10-12), with the most associated being position beta74. Our results are consistent with the possibility that position beta74 of the DRB1 molecule may have a specific and central role in autoantigen presentation by DRB1 to T lymphocytes. However, they also highlight the fact that it is not yet possible to rule out a role for DQA1.