title: Optimized detection of circulating anti-nuclear envelope autoantibodies by immunofluorescence.
abstract: BACKGROUND
  Antinuclear antibodies are useful diagnostic tools in several autoimmune diseases. However, the routine detection of nuclear envelope autoantibodies using immunofluorescence (IF) is not always easy to perform in patients' sera because of the presence of autoantibodies to other nuclear and cytoplasmic components which could mask the characteristic rim-like pattern of nuclear envelope autoantibodies. This is particularly common in sera from patients with primary biliary cirrhosis (PBC), which generaly have high titres of anti-mitochondrial antibodies. Therefore, we have assayed a number of commercial slides and alternative fixation conditions to optimize the detection of anti-nuclear envelope antibodies (ANEA) in PBC sera.
  
  
  METHODS
  We have explored the presence of ANEA in 33 sera from patients with established PBC using three different Hep2 commercial slides and home-made slides with HeLa and Hep2 cells fixed with methanol, ethanol, 1% or 4% formaldehyde.
  
  
  RESULTS
  We observed that the IF pattern was related to the cell type used (Hep2 or HeLa), the manufacturer and the cell fixation scheme. When both cell lines were fixed with 1% formaldehyde, the intensity of the cytoplasmic staining was considerably decreased regardless to the serum sample, whereas the prevalence of cytoplasmic autoantibodies was significantly lowered, as compared to any of the Hep2 commercial slide and fixation used. In addition, the prevalence of ANEA was importantly increased in formaldehyde-fixed cells.
  
  
  CONCLUSIONS
  Immunofluorescence using appropriately fixed cells represent an easy, no time-consuming and low cost technique for the routine screening of sera for ANEA. Detection of ANEA is shown to be more efficient using formaldehyde-fixed cells instead of commercially available Hep2 cells.
authors.affiliation: Biochemistry, University of Crete, School of Medicine, P,O, Box 2208, Heraklion 71003, Greece. bio1020@edu.biology.uoc.gr
authors.forename: Vagia
authors.initials: V
authors.lastname: Tsiakalou
authors.forename: Elena
authors.initials: E
authors.lastname: Tsangaridou
authors.forename: Hara
authors.initials: H
authors.lastname: Polioudaki
authors.forename: Artemissia-Phoebe
authors.initials: AP
authors.lastname: Nifli
authors.forename: Meri
authors.initials: M
authors.lastname: Koulentaki
authors.forename: Tonia
authors.initials: T
authors.lastname: Akoumianaki
authors.forename: Elias
authors.initials: E
authors.lastname: Kouroumalis
authors.forename: Elias
authors.initials: E
authors.lastname: Castanas
authors.forename: Panayiotis A
authors.initials: PA
authors.lastname: Theodoropoulos
mesh_terms: D000328:Adult; D000368:Aged; D000974:Antibodies, Antinuclear / Q000097:blood*; D045744:Cell Line, Tumor; D005260:Female; D005404:Fixatives; D019084:Fluorescent Antibody Technique, Indirect / Q000379:methods*; D005557:Formaldehyde; D006367:HeLa Cells; D006801:Humans; D008297:Male; D008856:Microscopy, Fluorescence; D008875:Middle Aged; D009685:Nuclear Envelope / Q000276:immunology*
chemical_list: D000974:Antibodies, Antinuclear; D005404:Fixatives; D005557:Formaldehyde