title: A dual-fixed neutrophil substrate improves interpretation of antineutrophil cytoplasmic antibodies by indirect immunofluorescence.
abstract: OBJECTIVE
  To determine whether the addition of a formalin-fixed neutrophil substrate could improve interpretation and prediction of autoantigenic specificity in antineutrophil cytoplasmic antibody (ANCA) testing.
  
  
  METHODS
  Routine diagnostic samples sent for ANCA testing were analyzed prospectively on a dual substrate of both ethanol- and formalin-fixed neutrophils. Positive samples on ethanol-fixed neutrophils were deemed "typical" if formalin-fixed neutrophils also stained, and "atypical" if not. Indirect immunofluorescence (IIF) results were correlated with antimyeloperoxidase (MPO) and anti-proteinase 3 (PR3) results with an enzyme-linked immunosorbent assay (ELISA).
  
  
  RESULTS
  Of 1,426 samples, 201 from unique patients were ANCA-positive (200 on IIF, 1 on ELISA alone). Thirty-two (45%) of 71 typical ANCA staining patterns were positive for either an anti-MPO or anti-PR3 antibodies, whereas only one (0.8%) of 129 atypical patterns was ELISA-positive, in a patient without systemic vasculitis. Only one (3%) of 34 ELISA-positive samples had a negative IIF-ANCA (1/1,426 patients, 0.07%), and this patient did not have vasculitis.
  
  
  CONCLUSIONS
  Concomitant staining on formalin fixation of IIF-positive ethanol-fixed ANCA samples improves the interpretation of ANCA testing and is predictive of vasculitis autoantigens MPO and PR3.
authors.affiliation: From the Department of Immunopathology, Pathology West, ICPMR, Westmead Hospital, Westmead, Australia, and Discipline of Medicine, Sydney Medical School, University of Sydney, Sydney, Australia. ming-wei.lin@sydney.edu.au.
authors.forename: Ming-Wei
authors.initials: MW
authors.lastname: Lin
authors.affiliation: From the Department of Immunopathology, Pathology West, ICPMR, Westmead Hospital, Westmead, Australia, and.
authors.forename: Roger A
authors.initials: RA
authors.lastname: Silvestrini
authors.affiliation: From the Department of Immunopathology, Pathology West, ICPMR, Westmead Hospital, Westmead, Australia, and.
authors.forename: Suzanne
authors.initials: S
authors.lastname: Culican
authors.affiliation: From the Department of Immunopathology, Pathology West, ICPMR, Westmead Hospital, Westmead, Australia, and.
authors.forename: David
authors.initials: D
authors.lastname: Campbell
authors.affiliation: From the Department of Immunopathology, Pathology West, ICPMR, Westmead Hospital, Westmead, Australia, and Discipline of Medicine, Sydney Medical School, University of Sydney, Sydney, Australia.
authors.forename: David A
authors.initials: DA
authors.lastname: Fulcher
mesh_terms: D056648:Anti-Neutrophil Cytoplasmic Antibody-Associated Vasculitis / Q000175:diagnosis* / Q000276:immunology; D019268:Antibodies, Antineutrophil Cytoplasmic / Q000032:analysis*; D000431:Ethanol; D019084:Fluorescent Antibody Technique, Indirect / Q000379:methods*; D005557:Formaldehyde; D006801:Humans; D009504:Neutrophils / Q000276:immunology*
chemical_list: D019268:Antibodies, Antineutrophil Cytoplasmic; D005557:Formaldehyde; D000431:Ethanol
keywords: Antineutrophil cytoplasmic antibody (ANCA); Formalin fixation; Indirect immunofluorescence (IIF); Vasculitis