Autoantibodies against neutrophil granulocytesare frequently observed in patients with ulcerativecolitis, but a precise description of the autoantigensinvolved is lacking. Therefore, sera from 75 patients with ulcerative colitis were studied forantibody specificities by means of indirectimmunofluorescence microscopy, enzyme-linkedimmunosorbent assays, and immunoblotting of extracts ofneutrophils, neutrophil granules and lymphocytes. Fifty-six percent ofthe sera reacted in indirect immunofluorescence withethanol-fixed neutrophils. On formalin-fixed cells mostsera shifted fluorescence pattern, indicating a cytoplasmic origin of antigen(s). Only a fewsera reacted with specific, known antigens. Immunoblotsshowed reactions with a broad panel of antigens withpreferences for proteins around 55-65 kDa, which were present in azurophilic granules and incytosol, an 80-kDa protein found in the specificgranules, and a 110-kDa unknown cytosol component. Inconclusion, neutrophilspecific IgG autoantibodies from ulcerative colitis patients react with severaldifferent antigens, most of which are of nonnuclearorigin.