Differentiation of antineutrophil nuclear antibodies in inflammatory boweland autoimmune liver diseases from antineutrophil cytoplasmic antibodies (p-ANCA) using immunofluorescence microscopy

Perinuclear antineutrophil cytoplasmic antibodies (p-ANCA) directed against cytoplasmic proteins of neutrophils have been studied extensively in patie nts with systemic vasculitides. Recent data indicate that antineutrophil an tibodies in sera from patients with chronic inflammatory bowel diseases (IB D) or autoimmune liver disorders, currently called 'atypical p-ANCA', recog nize a nuclear target antigen, rendering the term 'ANCA' inaccurate. Specif ic microscopic criteria to distinguish atypical p-ANCA from p-ANCA are lack ing. We used planar and confocal laser scanning indirect immunofluorescence microscopy to examine the labelling characteristics of ethanol-, methanol- and formaldehyde-fixed neutrophils by antineutrophil antibodies in 153 ser um samples from patients with IBD, autoimmune liver disorders, systemic vas culitides or healthy blood donors. On ethanol- or methanol-fixed neutrophil s, multiple intranuclear fluorescent foci together with either a rim-like p eripheral nuclear staining ('type A') or a combined cytoplasmic and periphe ral nuclear staining ('type B') was noted exclusively with atypical p-ANCA in sera from patients with IBD or autoimmune liver disorders. Intranuclear foci, which probably corresponded to invaginations of the nuclear envelope, were not labelled by p-ANCA from patients with microscopic polyangiitis or cytoplasmic ANCA (c-ANCA) from patients with Wegener's granulomatosis. On formaldehyde-fixed neutrophils, atypical p-ANCA gave a fine rim-like staini ng of the nuclear periphery, whereas ANCA diffusely labelled the cytoplasm. To distinguish reliably between the patterns produced by atypical p-ANCA o r p-ANCA, particularly p-ANCA, careful indirect immunofluorescence microsco py on ethanol- as well as on formaldehyde-fixed neutrophils is necessary, w ith particular emphasis on the presence of multiple intranuclear fluorescen t foci.