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Submitted on March 10, 2003
Revised on August 25, 2003
Accepted on September 29, 2003

A non-redundant human protein chip for antibody screening and serum profiling

Angelika Lueking, Alexandra Possling, Otmar Huber, Allan Beveridge, Martin Horn, Holger Eickhoff, Johannes Schuchardt, Hans Lehrach, and Dolores J. Cahill

Centre for Human Proteomics, Royal College of Surgeons in Ireland, Dublin 2

Corresponding Author: chp{at}rcsi.ie

There is burgeoning interest in protein microarrays, but a source of thousands of non-redundant, purified proteins was not previously available. Here we show a glass chip containing 2,413 non-redundant purified human fusion proteins on a polymer surface, where densities up to 1600 proteins/cm2 on a microscope slide can be realised. In addition, the polymer coating of the glass slide enables screening of protein interactions under non-denaturing conditions. Such screenings require only two hundred microlitres sample volumes, illustrating their potential for high-throughput applications. Here we demonstrate two applications, (i) the characterization of antibody binding, specificity and cross-reactivity and (ii) profiling the antibody repertoire in body fluids, such as serum from patients with auto-immune diseases. For (i), we have incubated these protein chips with anti-RGSHis6, anti-GAPDH, and anti-HSP90b antibodies. In an initial proof of principle study for (ii), we have screened serum from alopecia and arthritis patients. With analysis of large sample numbers, identification of disease-associated proteins to generate novel diagnostic markers may be possible.


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