ANCA Diagnostics in Clinical Practice: New Developments

Anti-neutrophil cytoplasmic antibodies (ANCA) are the hallmark of the so-called ANCAassociated vasculitides (AAV) (Jennette & Falk, 1997). These primary small-vessel vasculitides comprise granulomatosis with polyangiitis (GPA, previously referred to as Wegener’s granulomatosis) (Falk et al, 2011), the Churg-Straus syndrome (CSS), microscopic polyangiitis (MPA), and renal-limited necrotizing crescentic glomerulonephritis (NCGN). These disease entities can be discriminated based on systemic symptoms of disease (GPA, CSS, and MPA) versus clinical manifestations that are restricted to the kidneys (NCGN). The systemic diseases can further be subdivided by the presence (GPA and CSS) or absence (MPA) of granulomatous lesions in the airways, while GPA and CSS can be distinguished based on the presence of asthmatic manifestations and/or eosinophilia in CSS, but not in GPA. Classification criteria for these diseases have been defined by the American college of rheumatology (ACR; Fries et al, 1990) and the Chapel Hill consensus conference (Jennette, 1994). The presence of ANCA, however, is not part of these criteria which are primarily based on clinical manifestations and histopathology as observed in biopsies obtained from the affected tissues. More recently a novel consensus methodology for the classification of AAV was developed and validated for epidemiological studies (Watts et al, 2007). Importantly, the latter classification criteria incorporated the ANCA status of the patient. By definition ANCA are directed to the cytoplasmic components of neutrophilic granulocytes. In particular the constituents of the granules appear to be the antigenic targets. With respect to AAV the serine protease (PR)3 and myeloperoxidase (MPO) are the most important autoantigens. ANCA were originally detected by indirect immunofluorescence (IIF) on ethanol-fixed neutrophils. According to the international consensus on ANCA detection four patterns have to be distinguished (Savige et al, 1999). First, the classical (C-)ANCA is characterized by a granular, cytoplasmic fluorescence with central or interlobular accentuation; second, a diffuse flat cytoplasmic fluorescence without interlobular accentuation may be referred to as atypical C-ANCA. In clinical practice, however, this pattern is not distinguished by many clinical laboratories. Third, the perinuclear (P-)ANCA is characterized by perinuclear staining, with or without nuclear extension. Reading of the P-ANCA pattern may be hampered by the presence of interfering antinuclear antibodies (ANA). Finally, if a combination of cytoplasmic and perinuclear staining occurs, this is called atypical ANCA. The perinuclear staining pattern actually is an artefact, since formalin-fixation produces a cytoplasmic staining pattern, indistinguishable