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Two-color, rolling-circle amplification on antibody microarrays for sensitive, multiplexed serum-protein measurements

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Addresses: *The Van Andel Research Institute, 333 Bostwick, Grand Rapids, MI 49503, USA. †The University of Amsterdam, Department of Molecular and Cell Biology, 1081 BT Amsterdam, The Netherlands. ‡The University of Michigan Medical School, 1500 East Medical Center Drive, Ann Arbor, MI 48109, USA. §Yale University School of Medicine, Department of Pathology, 310 Cedar Street, New Haven, CT 06510, USA. Correspondence: Brian B Haab. E-mail: Brian.Haab@vai.org reviews Published: 30 March 2004 Genome Biology 2004, 5:R28 The electronic version of this article is the complete one and can be found online at. | Method Open Access Two-color rolling-circle amplification on antibody microarrays for sensitive multiplexed serum-protein measurements Heping Zhou Kerri Bouwman Mark Schotanus Cornelius Verweij Jorge A Marrero Deborah Dillon Jose Costa Paul Lizardi and Brian B Haab Addresses The Van Andel Research Institute 333 Bostwick Grand Rapids MI 49503 USA. The University of Amsterdam Department of Molecular and Cell Biology 1081 BT Amsterdam The Netherlands. The University of Michigan Medical School 1500 East Medical Center Drive Ann Arbor MI 48109 USA. Yale University School of Medicine Department of Pathology 310 Cedar Street New Haven CT 06510 USA. Correspondence Brian B Haab. E-mail Brian.Haab@vai.org Published 30 March 2004 Genome Biology 2004 5 R28 The electronic version of this article is the complete one and can be found online at http genomebiology.com 2004 5M R28 Received 11 November 2003 Revised 8 January 2004 Accepted 13 February 2004 2004 Zhou et al. licensee BioMed Central Ltd. This is an Open Access article verbatim copying and redistribution of this article are permitted in all media for any purpose provided this notice is preserved along with the article s original URL. Abstract The ability to conveniently and rapidly profile a diverse set of proteins has valuable applications. In a step toward further enabling such a capability we developed the use of rolling-circle amplification RCA to measure the relative levels of proteins from two serum samples labeled with biotin and digoxigenin respectively that have been captured on antibody microarrays. Two-color RCA produced fluorescence up to 30-fold higher than direct-labeling and indirect-detection methods using antibody microarrays prepared on both polyacrylamide-based hydrogels and nitrocellulose. Replicate RCA measurements of multiple proteins from sets of 24 serum samples were highly reproducible and accurate. In addition RCA enabled reproducible measurements of distinct expression profiles from .