Multiplex Imaging of Rho Family GTPase Activities in Living Cells
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Here, we provide procedures for imaging the Rho GTPase biosensors in both single and multiplex acquisition modes. The multiplex approach enables the direct visualization of two biosensor readouts from a single living cell. Here, we take as an example a combination of the RhoA biosensor based on a CFP/YFP FRET modality and the Cdc42 biosensor based on organic dyes that change fluorescence as a function of the local solvent polarity. We list the required optical components as well as cellular manipulation techniques necessary to successfully image these two ratiometric biosensors in a single living cell.
