Abstract

<h2>METHODS FOR DETECTION AND ANTIGENIC TARGETS OF ANCA</h2> Davies et al. ( 9 ) were the first to report the presence of antibodies that produced diffuse cytoplasmic staining of neutrophils by indirect immunofluorescence (IIF) techniques in patients with segmental necrotizing glomerulonephritis (GN). Several years later, the same antibody reaction pattern was recognized to occur in sera of patients with Wegener’s granulomatosis (WG) ( 63 ). A neutrophil perinuclear cytoplasmic staining pattern was subsequently noted to be associated with antibody reactivity in patients with microscopic polyangiitis (MPA) and idiopathic crescentic necrotizing GN ( 12 ). This led to a large number of studies that have examined the presence and pattern of ANCA in various disease states. Routinely, ANCA are detected by IIF on ethanol-fixed neutrophils ( 30 ). According to guidelines established at the 1st workshop on ANCA in 1988, neutrophils isolated from heparinized blood are cytocentrifuged, fixed in absolute ethanol on glass slides, and then incubated with dilutions of patient’s serum ( 67 ). Slides are stained with fluorescein-labeled anti-human immunoglobulin, and the fluorescence pattern is read with a fluorescence microscope ( 66 , 67 ). With this technique three distinct patterns of IIF can be observed (see