title: Miscellaneous vitreous-derived IgM antibodies target numerous retinal proteins in equine recurrent uveitis.
abstract: OBJECTIVE
  In equine recurrent uveitis (ERU), immune reactions are directed toward known antigens like S-antigen, interphotoreceptor retinoid-binding protein, and cellular retinalaldehyde-binding protein, and anti-retinal antibodies were detected in vitreous samples. The aim of this study was the investigation of intraocular immunoglobulin M (IgM) reactivities to retinal proteome.
  
  
  METHODS
  Retina was separated by one- and two-dimensional gel electrophoresis and blotted semidry on PVDF membranes. To identify intraocular IgM antibody responses to retinal tissue, blots were incubated with vitreous samples of ERU-diseased horses (n = 50) and healthy controls (n = 30), followed by an HRP-labeled secondary antibody specific for equine IgM. Noticeable 2D western blot signals were aligned on a 2D gel of retinal proteome, excised, and subsequently identified by tandem mass spectrometry.
  
  
  RESULTS
  Interestingly, frequent and very miscellaneous IgM response patterns to the retinal proteome in 68% of ERU vitreous samples were detected. Binding of IgM antibodies was localized at 17 different molecular weights. The most frequently detected signal, in 21 of the 50 samples, was located at 49 kDa. Comparing the samples interindividually between one and up to nine different signals in one sample could be observed. All healthy vitreous samples were devoid of IgM antibodies. Analysis of targeted spots with mass spectrometry led to the clear identification of 11 different proteins (corresponding to 16 different spots). One candidate could not be discovered so far.
  
  
  CONCLUSIONS
  The considerable IgM response to retinal proteins demonstrates an ongoing immune response, which might contribute to the remitting relapsing character of ERU. Novel identified target proteins point to a diverse response pattern of individual ERU cases.
authors.affiliation: Department of Veterinary Sciences, Institute of Animal Physiology, Ludwig-Maximilians University, Veterinärstr 13, D-80539 München, Germany.
authors.forename: Johanna K
authors.initials: JK
authors.lastname: Zipplies
authors.forename: Stefanie M
authors.initials: SM
authors.lastname: Hauck
authors.forename: Christina
authors.initials: C
authors.lastname: Eberhardt
authors.forename: Sieglinde
authors.initials: S
authors.lastname: Hirmer
authors.forename: Barbara
authors.initials: B
authors.lastname: Amann
authors.forename: Manfred
authors.initials: M
authors.lastname: Stangassinger
authors.forename: Marius
authors.initials: M
authors.lastname: Ueffing
authors.forename: Cornelia A
authors.initials: CA
authors.lastname: Deeg
mesh_terms: D000818:Animals; D001323:Autoantibodies; D005136:Eye Proteins / Q000276:immunology*; D006734:Horse Diseases / Q000276:immunology*; D006736:Horses; D007075:Immunoglobulin M / Q000276:immunology*; D020543:Proteome; D012160:Retina / Q000276:immunology; D014605:Uveitis / Q000276:immunology / Q000473:pathology / Q000662:veterinary*
chemical_list: D001323:Autoantibodies; D005136:Eye Proteins; D007075:Immunoglobulin M; D020543:Proteome