Protein Arrays III: Reverse-Phase Protein Arrays
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The reverse-phase protein array (RPPA) is to use highly specific antibodies to interrogate pan or posttranslationally modified protein targets, such as phosphorylated proteins, particularly the proteins involved in cell signaling pathways. In this protocol we will cover the preparation of cell (or tissue) lysates, sample printing, antibody validation, antibody interrogation, signal amplification steps, imaging and data analysis. In this protocol, colorimetric catalyzed signal amplification (CSA) chemistry, fluorescence and near-infrared (NIR) based detection methods will be described.
Key wordsReverse-phase protein array Antibody validation Cell signaling pathways Near-infrared (NIR) fluorescence
This work was partly supported by a grant from the Lupus Research Institute to T.W. and a startup fund from the University of Houston to T.W.
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