Characterization and epitope mapping of human monoclonal antibodies to PDC-E2, the immunodominant autoantigen of primary biliary cirrhosis

Patrick S C Leung, Sheri Krams, Santiago Munoz, Charles P. Surh, Aftab Ansari, Thomas Kenny, Dick L. Robbins, John Fung, Thomas E. Starzl, Willis Maddrey, Ross L. Coppel, M. Eric Gershwin

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Abstract

Further to define the epitopes of PDC-E2, the major autoantigen in primary biliary cirrhosis (PBC), we have developed and characterized five human monoclonal antibodies. These antibodies were derived by fusing a regional hepatic lymph node from a patient with PBC with the mouse human heterohybrid cell line F3B6. Previous studies of epitope mapping of PDC-E2 have relied on whole sera and have suggested that the immunodominant epitope lies within the inner lipoyl domain of the molecule. However, selective absorption studies using whole sera and a series of overlapping recombinant peptides of PDC-E2 have suggested that the epitope may also include a large conformational component. Moreover, several laboratories have suggested that autoantibodies against the 2-oxo acids dehydrogenase autoantigens are cross-reactive. The five monoclonal antibodies generated included three IgG2a and two IgM antibodies and were studied for antigen specificity using recombinant PDC-E2, recombinant BCKD-E2, histone, dsDNA, IgG (Fc), collagen and a recombinant irrelevant liver specific control, the F alloantigen. The antibodies were also used to probe blots of human, bovine, mouse and rat mitochondria. Finally, fine specificity was studied by selective ELISA and absorption against overlapping expressing fragments of PDC-E2. All five monoclonals, but none of the other mitochondrial autoantigens were specific for PDC-E2. In fact, although affinity purified antibodies to PDC-E2 from patients with PBC cross-reacted with protein X, the human monoclonals did not, suggesting that protein X contains an epitope distinct from that found on PDC-E2. Additionally, all three IgG2 monoclonals recognized distinct epitopes within the inner lipoyl domain of PDC-E2.

Original languageEnglish (US)
Pages (from-to)703-718
Number of pages16
JournalJournal of Autoimmunity
Volume5
Issue number6
DOIs
StatePublished - 1992

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Epitope Mapping
Biliary Liver Cirrhosis
Autoantigens
Epitopes
Monoclonal Antibodies
Antibodies
Immunoglobulin G
Keto Acids
Immunodominant Epitopes
Antibody Affinity
Isoantigens
Liver
Serum
Autoantibodies
Histones
Immunoglobulin M
Mitochondria
Oxidoreductases
Proteins
Collagen

ASJC Scopus subject areas

  • Immunology
  • Immunology and Allergy

Cite this

Characterization and epitope mapping of human monoclonal antibodies to PDC-E2, the immunodominant autoantigen of primary biliary cirrhosis. / Leung, Patrick S C; Krams, Sheri; Munoz, Santiago; Surh, Charles P.; Ansari, Aftab; Kenny, Thomas; Robbins, Dick L.; Fung, John; Starzl, Thomas E.; Maddrey, Willis; Coppel, Ross L.; Gershwin, M. Eric.

In: Journal of Autoimmunity, Vol. 5, No. 6, 1992, p. 703-718.

Research output: Contribution to journalArticle

Leung, PSC, Krams, S, Munoz, S, Surh, CP, Ansari, A, Kenny, T, Robbins, DL, Fung, J, Starzl, TE, Maddrey, W, Coppel, RL & Gershwin, ME 1992, 'Characterization and epitope mapping of human monoclonal antibodies to PDC-E2, the immunodominant autoantigen of primary biliary cirrhosis', Journal of Autoimmunity, vol. 5, no. 6, pp. 703-718. https://doi.org/10.1016/0896-8411(92)90187-U
Leung, Patrick S C ; Krams, Sheri ; Munoz, Santiago ; Surh, Charles P. ; Ansari, Aftab ; Kenny, Thomas ; Robbins, Dick L. ; Fung, John ; Starzl, Thomas E. ; Maddrey, Willis ; Coppel, Ross L. ; Gershwin, M. Eric. / Characterization and epitope mapping of human monoclonal antibodies to PDC-E2, the immunodominant autoantigen of primary biliary cirrhosis. In: Journal of Autoimmunity. 1992 ; Vol. 5, No. 6. pp. 703-718.
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abstract = "Further to define the epitopes of PDC-E2, the major autoantigen in primary biliary cirrhosis (PBC), we have developed and characterized five human monoclonal antibodies. These antibodies were derived by fusing a regional hepatic lymph node from a patient with PBC with the mouse human heterohybrid cell line F3B6. Previous studies of epitope mapping of PDC-E2 have relied on whole sera and have suggested that the immunodominant epitope lies within the inner lipoyl domain of the molecule. However, selective absorption studies using whole sera and a series of overlapping recombinant peptides of PDC-E2 have suggested that the epitope may also include a large conformational component. Moreover, several laboratories have suggested that autoantibodies against the 2-oxo acids dehydrogenase autoantigens are cross-reactive. The five monoclonal antibodies generated included three IgG2a and two IgM antibodies and were studied for antigen specificity using recombinant PDC-E2, recombinant BCKD-E2, histone, dsDNA, IgG (Fc), collagen and a recombinant irrelevant liver specific control, the F alloantigen. The antibodies were also used to probe blots of human, bovine, mouse and rat mitochondria. Finally, fine specificity was studied by selective ELISA and absorption against overlapping expressing fragments of PDC-E2. All five monoclonals, but none of the other mitochondrial autoantigens were specific for PDC-E2. In fact, although affinity purified antibodies to PDC-E2 from patients with PBC cross-reacted with protein X, the human monoclonals did not, suggesting that protein X contains an epitope distinct from that found on PDC-E2. Additionally, all three IgG2 monoclonals recognized distinct epitopes within the inner lipoyl domain of PDC-E2.",
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AU - Leung, Patrick S C

AU - Krams, Sheri

AU - Munoz, Santiago

AU - Surh, Charles P.

AU - Ansari, Aftab

AU - Kenny, Thomas

AU - Robbins, Dick L.

AU - Fung, John

AU - Starzl, Thomas E.

AU - Maddrey, Willis

AU - Coppel, Ross L.

AU - Gershwin, M. Eric

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AB - Further to define the epitopes of PDC-E2, the major autoantigen in primary biliary cirrhosis (PBC), we have developed and characterized five human monoclonal antibodies. These antibodies were derived by fusing a regional hepatic lymph node from a patient with PBC with the mouse human heterohybrid cell line F3B6. Previous studies of epitope mapping of PDC-E2 have relied on whole sera and have suggested that the immunodominant epitope lies within the inner lipoyl domain of the molecule. However, selective absorption studies using whole sera and a series of overlapping recombinant peptides of PDC-E2 have suggested that the epitope may also include a large conformational component. Moreover, several laboratories have suggested that autoantibodies against the 2-oxo acids dehydrogenase autoantigens are cross-reactive. The five monoclonal antibodies generated included three IgG2a and two IgM antibodies and were studied for antigen specificity using recombinant PDC-E2, recombinant BCKD-E2, histone, dsDNA, IgG (Fc), collagen and a recombinant irrelevant liver specific control, the F alloantigen. The antibodies were also used to probe blots of human, bovine, mouse and rat mitochondria. Finally, fine specificity was studied by selective ELISA and absorption against overlapping expressing fragments of PDC-E2. All five monoclonals, but none of the other mitochondrial autoantigens were specific for PDC-E2. In fact, although affinity purified antibodies to PDC-E2 from patients with PBC cross-reacted with protein X, the human monoclonals did not, suggesting that protein X contains an epitope distinct from that found on PDC-E2. Additionally, all three IgG2 monoclonals recognized distinct epitopes within the inner lipoyl domain of PDC-E2.

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