Quantitative label-free imaging of membrane protein interaction kinetics on cells Quantitative label-free imaging of membrane protein interaction kinetics on cells ABSTRACT Proteins embedded in or attached to cell membranes perform many critically important biological functions, including cell signaling, communication and the transport of vital substances into and out of cells. They are also the most important drug targets and disease biomarkers. Despite the importance, studying membrane proteins, especially quantifying their interactions with other molecules, such as drug candidates, has been a difficult challenge. Current methods rely on either labeling the proteins with fluorescent tags or extracting them from their native membrane environment, and then purifying and immobilizing them on a surface for binding kinetic studies. The former approach is an end-point-assay, which does not provide kinetics information required for quantifying protein interactions, while the latter is not only labor-intensive but also prone to alternation of the native structures and functions of the membrane proteins. This project focuses on developing a novel technique for studying and quantifying membrane protein interactions in their native cellular environment without the need of extraction, purification, or immobilization. The core of the technique is plasmonic-based electrical impedance microscopy (P-EIM) recently invented in the PIs lab, which has several unique capabilities: 1) It is label free and can provide quantitative analysis of binding kinetics; 2) It has a high spatial resolution (sub-microns), and thus is suitable for analyzing membrane protein binding activities of single cells, and for mapping local binding kinetics of membrane proteins within a single cell; 3) It is fast (ms), which enables real-time tracking of cell signal transduction cascade triggered by small molecule binding to membrane proteins. Additionally, the technique allows for simultaneous plasmonic, impedance and fluorescence imaging, and combines the strengths of these methods in one system.
|Effective start/end date||7/1/14 → 8/31/18|
- HHS-NIH: National Institute of General Medical Sciences (NIGMS): $1,125,670.00
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