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On silico peptide microarrays for high-resolution mapping of antibody epitopes and diverse protein-protein interactions

Nature Medicine volume 18, pages 1434–1440 (2012)Cite this article

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Abstract

We developed a new, silicon-based peptide array for a broad range of biological applications, including potential development as a real-time point-of-care platform. We used photolithography on silicon wafers to synthesize microarrays (Intel arrays) that contained every possible overlapping peptide within a linear protein sequence covering the N-terminal tail of human histone H2B. These arrays also included peptides with acetylated and methylated lysine residues, reflecting post-translational modifications of H2B. We defined minimum binding epitopes for commercial antibodies recognizing the modified and unmodified H2B peptides. We further found that this platform is suitable for the highly sensitive characterization of methyltransferases and kinase substrates. The Intel arrays also revealed specific H2B epitopes that are recognized by autoantibodies in individuals with systemic lupus erythematosus who have elevated disease severity. By combining emerging nonfluorescence-based detection methods with an underlying integrated circuit, we are now poised to create a truly transformative proteomics platform with applications in bioscience, drug development and clinical diagnostics.

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Figure 1: Intel array design and construction.
Figure 2: Epitope mapping of polyclonal and monoclonal antibodies using H2B Intel arrays.
Figure 3: Detection of dimethlyated Intel array peptides and direct monomethylation of lysine-containing Intel array peptides by the PKMT SETD7.
Figure 4: Conventional spotted microarray showing increased reactivity to whole histones and post-translationally modified histone peptides in sera from individuals with IFN-high SLE.
Figure 5: Differential reactivity to histone H2B peptides in a cohort of individuals with SLE.

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Acknowledgements

We thank H. Lee, J. Zhu and the other members of the peptide array team at Biomedical Life Science at Digital Health Intel for development of the early peptide array process and G. Credo for help with the Online Methods section of the manuscript. We thank D. Hall, K.C. Garcia, S. Sidhu, J. Haddon, J. Jarrell and other members of our laboratories for helpful discussion. J.V.P. was funded by a US National Science Foundation Graduate Research Fellowship (NSF GRFP) and is supported by the Stanford Genome Training Program (SGTP; US National Institutes of Health (NIH), US National Human Genome Research Institute). D.L. is supported by the European Molecular Biology Organization, the Human Frontier Science Program and the Machiah Foundation. O.G. is a recipient of an Ellison Senior Scholar Award and is funded by a grant from the NIH (R01 GM079641). P.J.U. is the recipient of a Donald E. and Delia B. Baxter Foundation Career Development Award and is supported by National Heart, Lung, and Blood Institute (NHLBI) Proteomics contract HHSN288201000034C, Proteomics of Inflammatory Immunity and Pulmonary Arterial Hypertension; grants from the NIH (5 U19-AI082719, 5 U19-AI050864, 5 U19-AI056363, 1 U19 AI090019 and 4 U19 AI090019); Canadian Institutes of Health Research (2 OR-92141); Alliance for Lupus Research (grant number 21858); a gift from the Ben May Trust and a gift from the Floren Family Trust. The research leading to these results has received funding from the European Union Seventh Framework Programme (FP7/2007-2013) under grant agreement number 261382. C.L.L. is a recipient of an NIH National Research Service Award Fellowship (5 F32 AI-080086-02).

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Author notes

  1. Madoo Varma, Paul J Utz and Chih Long Liu: These authors contributed equally to this work.

  2. pjutz@stanford.edu

Authors and Affiliations

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

    Jordan V Price, Stephanie Tangsombatvisit, Paul J Utz & Chih Long Liu

  2. Intel Corporation, Santa Clara, California, USA

    Guangyu Xu, Jiangtao Yu & Madoo Varma

  3. Department of Biology, Stanford University, Stanford, California, USA

    Dan Levy & Or Gozani

  4. Center for Immunology, University of Minnesota Medical School, Minneapolis, Minnesota, USA

    Emily C Baechler

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Contributions

J.V.P. conducted antibody-binding experiments and ELISA, analyzed the data, made the figures and wrote the manuscript. S.T. performed antibody-binding experiments and contributed to data analysis and figure production. G.X. and J.Y. designed and fabricated Intel arrays. D.L. and O.G. contributed the methylation assay. E.C.B. characterized the IFN signature and provided ABCoN patient samples. M.V. supervised the development team at Intel and edited the manuscript. P.J.U. and C.L.L. supervised the project and edited the manuscript.

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Correspondence to Jordan V Price.

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M.V. and G.X. are employees of Intel.

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Price, J., Tangsombatvisit, S., Xu, G. et al. On silico peptide microarrays for high-resolution mapping of antibody epitopes and diverse protein-protein interactions. Nat Med 18, 1434–1440 (2012). https://doi.org/10.1038/nm.2913

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