Autoantibodies to neutrophil cytoplasmic antigens are found in patients with small or medium-size vessel systemic vasculitis such as Wegener's granulomatosis and microscopic polyarteritis. We studied the specificity of anti-neutrophil cytoplasm autoantibodies (ANCA) by examining the binding of sera from patients with different forms of systemic vasculitis, to neutrophil extract size-fractionated by gel filtration for high performance liquid chromatography. Sequential fractions were collected for solid-phase radioimmunoassay. Patients with Wegener's granulomatosis showed two peaks of activity over fractions of 12 and 2 kD. Patients with microscopic polyarteritis had two different peaks of activity over fractions of 100 and 25 kD. Patients with Takayasu's arteritis and dermal lymphocytic vasculitis were studied as controls (all were ANCA negative by indirect immunofluorescence studies and solid-phase radioimmunoassay). However, the patient with lymphocytic vasculitis showed a distinct peak of activity over 22 kD and patients with Takayasu's arteritis had a single peak of activity over 200 kD. These results suggest the binding pattern of sera to fractionated neutrophil extract could be useful in differentiating between different diseases within the spectrum of systemic vasculitis. Cryptic antigen recognition in certain vasculitides may only be revealed by this procedure. We conclude that the association of ANCA and systemic vasculitis may not be limited to diseases of small or medium-size vessels, but could also include large vessel disease such as Takayasu's arteritis.