title: Clinical and serological evaluation of a novel CENP-A peptide based ELISA.
abstract: BACKGROUND
  Anti-centromere antibodies (ACA) are useful biomarkers in the diagnosis of systemic sclerosis (SSc). ACA are found in 20 to 40% of SSc patients and, albeit with lower prevalence, in patients with other systemic autoimmune rheumatic diseases. Historically, ACA were detected by indirect immunofluorescence (IIF) on HEp-2 cells and confirmed by immunoassays using recombinant CENP-B. The objective of this study was to evaluate a novel CENP-A peptide ELISA.
  
  
  METHODS
  Sera collected from SSc patients (n=334) and various other diseases (n=619) and from healthy controls (n=175) were tested for anti-CENP-A antibodies by the novel CENP-A enzyme linked immunosorbent assay (ELISA). Furthermore, ACA were determined in the disease cohorts by IIF (ImmunoConcepts, Sacramento, CA, USA), CENP-B ELISA (Dr. Fooke), EliA CENP (Phadia, Freiburg, Germany) and line-immunoassay (LIA, Mikrogen, Neuried, Germany). Serological and clinical associations of anti-CENP-A with other autoantibodies were conducted in one participating centre. Inhibition experiments with either the CENP-A peptide or recombinant CENP-B were carried out to analyse the specificity of anti-CENP-A and -B antibodies.
  
  
  RESULTS
  The CENP-A ELISA results were in good agreement with other ACA detection methods. According to the kappa method, the qualitative agreements were: 0.73 (vs. IIF), 0.81 (vs. LIA), 0.86 (vs. CENP-B ELISA) and 0.97 (vs. EliA CENP). The quantitative comparison between CENP-A and CENP-B ELISA using 265 samples revealed a correlation value of rho=0.5 (by Spearman equation). The receiver operating characteristic analysis indicated that the discrimination between SSc patients (n=131) and various controls (n=134) was significantly better using the CENP-A as compared to CENP-B ELISA (P<0.0001). Modified Rodnan skin score was significantly lower in the CENP-A negative group compared to the positive patients (P=0.013). Inhibition experiments revealed no significant cross reactivity of anti-CENP-A and anti-CENP-B antibodies. Statistically relevant differences for gender ratio (P=0.0103), specific joint involvement (Jaccoud) (P=0.0006) and anti-phospholipid syndrome (P=0.0157) between ACA positive SLE patients and the entire SLE cohort were observed.
  
  
  CONCLUSIONS
  Anti-CENP-A antibodies as determined by peptide ELISA represent a sensitive, specific and independent marker for the detection of ACA and are useful biomarkers for the diagnosis of SSc. Our data suggest that anti-CENP-A antibodies are a more specific biomarker for SSc than antibodies to CENP-B. Furthers studies are required to verify these findings.
authors.affiliation: Dr. Fooke Laboratorien, Mainstrasse 85, 41469 Neuss, Germany.
authors.forename: Michael
authors.initials: M
authors.lastname: Mahler
authors.forename: Liesbeth
authors.initials: L
authors.lastname: Maes
authors.forename: Daniel
authors.initials: D
authors.lastname: Blockmans
authors.forename: Rene
authors.initials: R
authors.lastname: Westhovens
authors.forename: Xavier
authors.initials: X
authors.lastname: Bossuyt
authors.forename: Gabriela
authors.initials: G
authors.lastname: Riemekasten
authors.forename: Sandra
authors.initials: S
authors.lastname: Schneider
authors.forename: Falk
authors.initials: F
authors.lastname: Hiepe
authors.forename: Andreas
authors.initials: A
authors.lastname: Swart
authors.forename: Irmgard
authors.initials: I
authors.lastname: Gürtler
authors.forename: Karl
authors.initials: K
authors.lastname: Egerer
authors.forename: Margrit
authors.initials: M
authors.lastname: Fooke
authors.forename: Marvin J
authors.initials: MJ
authors.lastname: Fritzler
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D001323:Autoantibodies / Q000097:blood*; D001324:Autoantigens / Q000276:immunology*; D015415:Biomarkers / Q000097:blood; D016022:Case-Control Studies; D000076247:Centromere Protein A; D051936:Centromere Protein B / Q000276:immunology; D002868:Chromosomal Proteins, Non-Histone / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D012372:ROC Curve; D012595:Scleroderma, Systemic / Q000097:blood* / Q000276:immunology; D012680:Sensitivity and Specificity; D055815:Young Adult
chemical_list: D001323:Autoantibodies; D001324:Autoantigens; D015415:Biomarkers; C000619700:CENPA protein, human; D000076247:Centromere Protein A; D051936:Centromere Protein B; D002868:Chromosomal Proteins, Non-Histone