Universal non-antibody detection of protein phosphorylation using pIMAGO.
This article describes methods for a new, non-antibody phosphorylation detection reagent, termed pIMAGO (phospho-imaging). This novel reagent takes advantage not only of the unique properties of the soluble nanoparticles, but also of the multiple functionalities of the molecule, allowing for highly selective, sensitive, and quantitative assessment of protein phosphorylation without using radioactive isotopes or phospho-specific antibodies. The methods allow for multiplexed detection of phosphorylation and total protein amount simultaneously. The straightforward and routine detection and quantitation of general phosphorylation on any site of any protein can be performed in western blot and ELISA formats.
Department of Biochemistry, Purdue University, West Lafayette, Indiana; Tymora Analytical Operations, LLC, West Lafayette, Indiana.
Anton B
AB
Iliuk
W Andy
WA
Tao
D001710:Biotin / Q000737:chemistry; D050091:Dendrimers / Q000737:chemistry; D057054:Molecular Imaging / Q000379:methods*; D010710:Phosphates / Q000737:chemistry; D010766:Phosphorylation; D011506:Proteins / Q000032:analysis* / Q000737:chemistry; D014025:Titanium / Q000737:chemistry
D050091:Dendrimers; D010710:Phosphates; D011506:Proteins; D001710:Biotin; D014025:Titanium
ELISA; high-throughput screening; kinase assay; phosphoprotein detection; phosphorylation; western blot