E-Book Overview
A diverse collection of unique methodologies to synthesize and construct protein arrays for basic and clinical research in a high-throughput manner. These readily reproducible techniques can be applied to such applications as quantifying specific proteins of interest and discovering novel proteins, as well as to a variety of affinity substances, including antibodies, peptides, proteins, aptamers, and chemicals. The techniques can be used for biomarkers discovery, assay development, clinical sample testing, and building the next generation of molecular tools.
E-Book Content
METHODS IN MOLECULAR BIOLOGY
TM TM
Volume 264
Protein Arrays Methods and Protocols Edited by
Eric T. Fung, MD, PhD
Protein Arrays From cDNA Expression Libraries
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1 Protein Arrays From cDNA Expression Libraries Hendrik Weiner, Thomas Faupel, and Konrad Büssow Summary This chapter describes the production of a cDNA expression library from human fetal brain, the construction of a high-density protein array from such a library, and two applications to screen the array for binding proteins. After producing the library and decollating the expression clones, one can pick thousands of expression clones with a laboratory robot and can deposit them into microtiter plates in an ordered manner. Such ordered clone libraries are the starting material for the construction of a high-density protein array. This array is constructed by spotting the expression clones onto a protein-binding membrane. Following cell growth and induction of protein expression on the membrane, the cell spots are lysed and their recombinant protein immobilized on the membrane. The so-constructed array carries thousands of proteins without the need to clone, express, and spot individual proteins. Such arrays allow one to screen for numerous protein functions in a high-throughput mann