Abstract: Objective To purify and quantify the autoantibodies against various epitopes within BP180-NC16A domain from the sera of patients with bullous pemphigoid （BP）. Methods Three epitopes within BP180-NC16A domain, NC16A-1, NC16A-2 and NC16A-3, were prepared. Blood samples were obtained from 10 patients who were diagnosed as active BP by clinical, pathological and immunofluorescence examination. Autoantibodies against these epitopes were purified with affinity chromatography column from the sera of these patients with active BP, and quantified separately. The relative binding affinity of autoantibodies to NC16A-1, NC16A-2 and NC16A-3 was measured using thiocyanate elution method. Results The autoantibodies against NC16A-1, NC16A-2 and NC16A-3 were successfully purified from the sera of patients. On average, the amount was 49.0 ± 20.7 μg, 117.7 ± 22.4 μg and 39.5 ± 18.9 μg respectively for autoantibodies to NC16A-1, NC16A-2 and NC16A-3 purified from a portion of serum containing about 20 mg IgG. Both the amount and binding affinity of anti-NC16A-2 autoantibody were significantly higher than those of anti-NC16A-1 and anti-NC16A-3 autoantibodies. Conclusion BP180 NC16A-2 （aa507-aa520） may be the major epitope recognized by pathogenic autoantibodies in patients with BP.

Key words: Bullous pemphigoid, Autoantibody