Familial clustering of Graves' disease indicates a genetic etiology. Searches for genetic factors additional to the known human leukocyte antigen (HLA) association have implicated the gene for the TSH receptor (TSHR). We analyzed the linkage and association among three recently described microsatellite markers within the TSHR introns in Graves' disease in large multiply affected Welsh and English families (223 members, 44 affected individuals). Linkage analysis under a dominant model strongly rejected the hypothesis that TSHR is linked to Graves' disease in these families (lod score = -4.53). More detailed analyses also failed to provide evidence for linkage; these included combined segregation and linkage analysis, correction for HLA-DR3 status, allowance for the levels of thyroid autoantibodies in unaffected pedigree members, consideration of a recessive model for the disease, and linkage disequilibrium between disease and marker alleles. We also considered the possibility of a genetic heterogeneity of Graves' disease and thus analyzed separately the different families with a similar result. Although these results cannot eliminate a minor role of the TSHR gene locus in the genetics of Graves' disease, they argue against it being a major genetic determinant in this pathology.