A dot-blot-staining method for detecting phosphoproteins with a Phos-tag Aqua fluorescent dye

• Kinoshita-Kikuta Emiko

  Department of Functional Molecular Science, Graduate School of Biomedical and Health Sciences, Hiroshima University

• Akayama Keisuke

  Department of Functional Molecular Science, Graduate School of Biomedical and Health Sciences, Hiroshima University

• Kinoshita Eiji

  Department of Functional Molecular Science, Graduate School of Biomedical and Health Sciences, Hiroshima University

• Koike Tohru

  Department of Functional Molecular Science, Graduate School of Biomedical and Health Sciences, Hiroshima University

Abstract

We describe a method for detecting phosphoproteins by dot-blot staining with a Phos-tag Aqua fluorescent dye. By using the method, three types of in vitro protein kinase assays for visualizing Tyr-, His-, and Asp-phosphoproteins, respectively, were performed. The staining procedure, which requires less than 2.5 h to complete, is conducted under conditions of neutral pH that are particularly advantageous for detecting labile His- and Asp-phosphoproteins. This method promises rapid and easy detection of phosphoproteins, and should be useful in high-throughput profiling of in vitro kinase activities or in vitro kinase inhibition.

Journal

• Journal of Electrophoresis

  Journal of Electrophoresis 64 (1), 7-11, 2020

  Japanese Electrophoresis Society

Keywords

• Phos-tag Aqua
• dot-blot staining
• tyrosine phosphorylation
• histidine phosphorylation
• aspartic acid phosphorylation

Details

• CRID: 1391694356246536704
• NII Article ID: 130007961315
• DOI: 10.2198/jelectroph.64.7
• ISSN: 13499408
• ISSN: 13499394
• Web Site: https://www.jstage.jst.go.jp/article/jelectroph/64/1/64_7/_pdf
• Web Site: https://search.jamas.or.jp/link/ui/2021342431
• Text Lang: en
• Data Source:
  • JaLC
  • Crossref
  • CiNii Articles
  • KAKEN
• Abstract License Flag: Disallowed