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Published on: September 19, 2018
Determining Protein Phosphorylation Status Using Antibody Arrays and Phos-Tag Biotin
Eiji Kinoshita1, Emiko Kinoshita-Kikuta2, Tohru Koike2
1Department of Functional Molecular Science, Institute of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan. kinoeiji@hiroshima-u.ac.jp.
Abstract:
We describe here a standard protocol for determining the phosphorylation status of protein multiplexes using antibody arrays and a biotinylated Phos-tag with a dodeca(ethylene glycol) spacer (Phos-tag Biotin). The procedure is based on an antibody microarray technique used in conjunction with an enhanced chemiluminescence system, and it permits the simultaneous and highly sensitive detection of multiple phosphoproteins in a cell lysate. By using this procedure, we have demonstrated the quantitative detection of the entire phosphorylation status of a target protein involved in intracellular signaling.

