Biosensors for characterizing the dynamics of rho family GTPases in living cells.

The biosensors developed in the authors' laboratory have been based on different designs, each imparting specific strengths and weaknesses. Here we describe detailed protocols for the application of three biosensors exemplifying different designs-first, a design in which an environmentally sensitive dye is used to report the activation of endogenous Cdc42, followed by two biosensors based on FRET, one using intramolecular and the other intermolecular FRET. The design differences lead to the need for different approaches in imaging and image analysis. (c) 2010 by John Wiley & Sons, Inc.