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Autoantigen microarrays for multiplex characterization of autoantibody responses

Nature Medicine volume 8, pages 295–301 (2002)Cite this article

Abstract

We constructed miniaturized autoantigen arrays to perform large-scale multiplex characterization of autoantibody responses directed against structurally diverse autoantigens, using submicroliter quantities of clinical samples. Autoantigen microarrays were produced by attaching hundreds of proteins, peptides and other biomolecules to the surface of derivatized glass slides using a robotic arrayer. Arrays were incubated with patient serum, and spectrally resolvable fluorescent labels were used to detect autoantibody binding to specific autoantigens on the array. We describe and characterize arrays containing the major autoantigens in eight distinct human autoimmune diseases, including systemic lupus erythematosus and rheumatoid arthritis. This represents the first report of application of such technology to multiple human disease sera, and will enable validated detection of antibodies recognizing autoantigens including proteins, peptides, enzyme complexes, ribonucleoprotein complexes, DNA and post-translationally modified antigens. Autoantigen microarrays represent a powerful tool to study the specificity and pathogenesis of autoantibody responses, and to identify and define relevant autoantigens in human autoimmune diseases.

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Figure 1: The 1152-feature rheumatic disease autoantigen array.
Figure 2: Array validation and sensitivity analysis.
Figure 3: Identification of disease-specific autoantibodies in human serum.
Figure 4: Autoantibody detection is specific.
Figure 5: Antigen arrays for epitope mapping, antibody isotype characterization, and comparative analysis.

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Acknowledgements

We thank K. Chong and D. Mitchell for their outstanding assistance; J.G. Lindsay for antibodies; and A. Abeliovich, H. Garren, P. Ruiz and G. Hermans for insightful discussions. This work was primarily supported by a Howard Hughes Postdoctoral Research Fellowship for Physicians Award, NIH K08 AR02133 and Arthritis Foundation Chapter Grant to W.H.R; NIH/NINDS 5R01NS18235 and NIH U19DK61934 to L.S.; and NIH K08 AI01521, an Arthritis Foundation Investigator Award, a Baxter Foundation Career Development Award, a Stanford University Office of Technology Licensing Award, and NIH U19 DK61934 to P.J.U. P.O.B is an Associate Investigator of the Howard Hughes Medical Institute.

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Authors and Affiliations

  1. Division of Immunology and Rheumatology, Department of Medicine, Stanford, California, USA

    William H. Robinson, Carla DiGennaro, Wolfgang Hueber, Makoto Kamachi, Erik J. Dean, Derek Fong, Mark C. Genovese, Henry E. Neuman De Vegvar & Paul J. Utz

  2. Department of Neurology, Stanford, California, USA

    William H. Robinson, Carla DiGennaro, Erik J. Dean, Derek Fong, David L. Hirschberg & Lawrence Steinman

  3. Department of Biochemistry and Howard Hughes Medical Institute, Stanford University School of Medicine, Stanford, California, USA

    Patrick O. Brown

  4. Tolerion, Palo Alto, California, USA

    William H. Robinson, Lawrence Steinman & Paul J. Utz

  5. Van Andel Research Institute, Grand Rapids, Michigan, USA

    Brian B. Haab

  6. Institut de Biologie Moléculaire et Cellulaire, CNRS, Strasbourg, France

    Sylvie Fournel & Sylviane Muller

  7. Division of Rheumatology, Department of Internal Medicine III, University of Vienna, Vienna, Austria

    Karl Skriner & Josef S. Smolen

  8. Rheumatology Diagnostics Laboratory, Los Angeles, California, USA

    Robert I. Morris

  9. Department of Biochemistry, Katholieke Universiteit Nijmegen, Nijmegen, the Netherlands

    Ger J. Pruijn & Walther J. van Venrooij

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  1. William H. Robinson
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  18. Patrick O. Brown
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  19. Lawrence Steinman
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  20. Paul J. Utz
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Correspondence to William H. Robinson.

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Robinson, W., DiGennaro, C., Hueber, W. et al. Autoantigen microarrays for multiplex characterization of autoantibody responses. Nat Med 8, 295–301 (2002). https://doi.org/10.1038/nm0302-295

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