Identification of rat targets of anti-soluble liver antigen autoantibodies by serologic proteome analysis.
BACKGROUND
Anti-soluble liver antigen (SLA) autoantibodies are specific for autoimmune hepatitis type 1 and are the only immunologic marker found in 15-20% of hepatitis cases previously considered cryptogenic. Anti-SLA antibodies react with the 100 000g supernatant from rat liver homogenate, but the molecular targets remain controversial.


METHODS
We characterized anti-SLA targets by one- and two-dimensional immunoblotting analysis. The recognized proteins were identified by peptide mass fingerprint analysis after matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry.


RESULTS
Three proteins of 35 kDa and pI 6.0, 50 kDa and pI between 6.0 and 6.5, and 58 kDa and pI between 6.5 and 7.0 were stained more intensely by anti-SLA positive-sera than by control sera. After in-gel tryptic digestion, MALDI-TOF analysis of the generated peptides enabled the clear identification of N-hydroxyarylamine sulfotransferase, isoforms of alpha-enolase, and isoforms of catalase.


CONCLUSIONS
Possible antigens for anti-SLA antibodies include a sulfotransferase, alpha-enolase(s), and catalase(s). Two-dimensional electrophoresis combined with mass spectrometry offers a versatile tool to identify molecular targets of autoantibodies and thus to improve diagnostic tools and the understanding of the immune process.
Ballot
Eric
E
Service d'Immunologie et Hématologie Biologiques, Hôpital Saint-Antoine, AP-HP, 75012 Paris, France.
Bruneel
Arnaud
A
Labas
Valérie
V
Johanet
Catherine
C
D000818:Animals; D001323:Autoantibodies / Q000097:blood*; D001324:Autoantigens / Q000276:immunology*; D015415:Biomarkers / Q000097:blood; D002374:Catalase / Q000097:blood* / Q000737:chemistry / Q000276:immunology; D015180:Electrophoresis, Gel, Two-Dimensional; D019084:Fluorescent Antibody Technique, Indirect; D019693:Hepatitis, Autoimmune / Q000097:blood / Q000276:immunology; D006801:Humans; D015151:Immunoblotting; D007527:Isoenzymes / Q000097:blood / Q000737:chemistry / Q000276:immunology; D008297:Male; D008970:Molecular Weight; D010449:Peptide Mapping; D010751:Phosphopyruvate Hydratase / Q000097:blood* / Q000737:chemistry / Q000276:immunology; D051381:Rats; D017208:Rats, Wistar; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D015238:Sulfotransferases / Q000097:blood* / Q000737:chemistry / Q000276:immunology
D001323:Autoantibodies; D001324:Autoantigens; D015415:Biomarkers; D007527:Isoenzymes; C046431:liver antigen LA-1; D002374:Catalase; C084221:St1C1 N-hydroxyarylamine sulfotransferase; D015238:Sulfotransferases; D010751:Phosphopyruvate Hydratase