Quantifying dimerization of receptor tyrosine kinase by quantum dot FRET Immunoassay in tissue sections

SummaryThis proposal is to develop a novel slide based proximity assay platform to quantify the dimerization of receptor tyrosine kinasesRTKin tissueQuantification of alteration of RTK dimerization upon biological stimuli or antagonist is critical to evaluate anti cancer therapies which aim to block RTK mediated growth signaling pathwayHoweverthere are limited methods for quantification of protein dimerization in tissue samplesRecentlywe have discovered that dual emission quantum dots can be used for Fluorescence Resonance Energy TransferFRETassaysin which one emission peak without FRET serves as an internal standard and the other serves as FRET emission to interact with acceptor dyeIn this proposalwe will use EGFR targeted therapies in squamous cell carcinoma of the head and neck as a model system to demonstrate the feasibility of using QD FRET for quantifying protein dimerization on tissuesThree specific aims are proposed in Phase I studyAimPrepare QD Ab conjugates with desired affinity and specificityAimDevelop quantification method for the QD FRET assayAimValidate QD FRET assay in human SCCHN cell lines and xenograft tissuesCompletion of this Phase I project will greatly facilitate further development of the QD FRET technology for both research and clinical settings Project NarrativeThis project aims to develop a quantum dot based immunoassay to quantify dimerization of receptor tyrosine kinases using proximity technology of Fluorescence Resonance Energy TransferThe developed novel proximity technology can improve biomarker evaluation using slide based immunofluorescent assay and have great research value and significant clinical impact for therapy management