Arrays of protein-capture agents and methods of use thereof

Abstract

Arrays of protein-capture agents useful for the simultaneous detection of a plurality of proteins which are the expression products, or fragments thereof, of a cell or population of cells in an organism are provided. A variety of antibody arrays, in particular, are described. Methods of both making and using the arrays of protein-capture agents are also disclosed. The invention arrays are particularly useful for various proteomics applications including assessing patterns of protein expression and modification in cells.

Description

[0001] This application is a continuation of co-pending application Ser. No. 09 / 574,748, filed May 18, 2000, which is a continuation-in-part of co-pending application Ser. No. 09 / 1115,455, filed Jul. 14, 1998, both of which are incorporated herein by reference in their entirety for all purposes and the specific purposes disclosed throughout this application.[0002] a) Field of the Invention[0003] The present invention relates generally to arrays of protein-capture agents and methods for the parallel detection and analysis of up to a large number of proteins in a sample. More specifically, the present invention relates to proteomics and the measurement of gene activity at the protein level in cells.[0004] b) Description of Related Art[0005] Although attempts to evaluate gene activity and to decipher biological processes including those of disease processes and drug effects have traditionally focused on genomices, proteomics offers a more direct and promising look a the biological func...

AI Technical Summary

Problems solved by technology

This 2D-gel technique requires large sample sizes, is time consuming, and is currently limited in its ability to reproducibly resolve a significant fraction of the proteins expressed by a human cell.

Techniques involving some large-format 2D-gels can produce gels which separate a larger number of proteins than traditional 2D-gel techniques, but reproducibility is still poor and over 95% of the spots cannot be sequenced due to limitations with respect to sensitivity of the available sequencing techniques.

The electrophoretic techniques are also plagued by a bias towards proteins of high abundance.

However, these multianalyte assays have not been directed towards assaying the total or partial protein content of a cell or cell population.

Furthermore, sample sizes required to adapt such standard antibody assay approaches to the analysis of even a fraction of the estimated 100,000 or more different proteins of a human cell and their various modified states are prohibitively large.

Materials, surface coatings, and detection methods used for macroscopic immunoassays and affinity purification are not readily transferable to the formation or fabrication of miniaturized protein arrays.

However, DNA biochip technology is not transferable to protein-binding assays such as antibody assays because the chemistries and materials used for DNA biochips are not readily transferable to use with proteins.

For instance, exposure of a substrates such as silicon, to air results in oxidation of the exposed surface.

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