Figure 2.

Optimization of RhoC activation assay. The G-LISA RhoA, B, and C absorbance-based GTPase activation assay was optimized for detecting active RhoC GTPase. Recombinant RhoC protein was labeled with nonhydrolyzable GTPγS, and 5 ng of protein was used per assay well. A 1:250 dilution of pan-Rho antibody was compared with decreasing antibody dilutions of a polyclonal RhoC-specific antibody, and a 1:250 dilution of an HRP-conjugated antirabbit secondary antibody. Absorbance was read at 490 nm, and data are presented as the mean ± SD of triplicate wells from a representative experiment (n = 3).