Patent ID: 7981633

Claim:
A method for measuring autophosphorylation of a tyrosine kinase in presence of a kinase inhibitor compared to autophosphorylation of said tyrosine kinase in absence of said kinase inhibitor, comprising (a) starving cells by serum depletion, (b) inducing kinase autophosphorylation activity by adding serum, growth factors and/or cytokines in presence and in absence of a kinase inhibitor, (c) solubilizing the cells to release a cell lysate, (d) capturing the tyrosine kinase in the cell lysate by adding a plurality of tyrosine kinase specific binding proteins, wherein each binding protein is associated with a unique dye, (e) adding a phosphotyrosine specific antibody tagged with a marker, wherein said marker is distinguishable from any of the unique dyes from d), (f) identifying the autophosphorylated tyrosine kinase by detecting the unique dye from d) and the marker from e), and (g) comparing a level of autophosphorylation of said tyrosine kinase in the presence of said kinase inhibitor to a level of autophosphorylation of said tyrosine kinase in the absence of said kinase inhibitor, wherein in case of autophosphorylation, a first signal for said unique dye and a second signal for said marker are detected whereas in case of lack of autophosphorylation, only the first signal for the unique dye is detected.