Four human monoclonal antibodies (SP1.4, WR1.7, TR1.8, and TR1.9) map the immunodominant region on thyroid peroxidase (TPO) recognized by autoantibodies in patients' sera. We used a pool of these monoclonal antibodies, expressed in bacteria as antigen-binding fragments [F(ab)], to compete for TPO autoantibody binding to radiolabeled TPO. The F(ab) inhibited TPO binding by 32 patients' sera by 82 +/- 14% (mean +/- SD), with a range from 51-100%. When each F(ab) was tested individually for its ability to compete for autoantibody binding to TPO, F(ab) TR1.8 was the most potent among the 32 sera. However, there was a wide spectrum of TPO binding inhibition when each serum was considered individually, thereby allowing an epitopic "fingerprint" to be drawn for the TPO autoantibodies in a patient's serum. There was a close association between the proportions of TPO autoantibodies to the TR1.8 and TR1.9 epitopes as well as between those to the SP1.4 and WR1.7 epitopes. These associations correspond to the previously described A and B epitopic domains in the TPO immunodominant region. No TPO epitope was observed to be associated with clinically apparent ophthalmopathy of Graves' disease, nor was there an association between TPO epitopes and patient age or sex. In summary, the present study on a large sample of sera with TPO autoantibodies indicates that by using TPO-specific F(ab) selected to cover all regions of the TPO immunodominant region, it is possible to obtain a TPO epitopic fingerprint for each serum. These data open the way to future studies directed at testing the hypothesis of disease-associated TPO epitope(s).