Flow cytometry was performed as described (18 (link)). Briefly, overnight E. coli cultures were diluted to OD600 = 0.1 and grown for 1 h at 37°C with shaking. Bacteria were then treated with 10 μM FITC-labeled TAT-RasGAP317-326. After 1 h of treatment, bacteria were washed with PBS and diluted 5-fold. For each sample, 10,000 events were then collected using a CytoFLEX benchtop flow cytometer (Beckman Coulter, Brea, CA). When indicated, extracellular fluorescence was quenched using 0.2% Trypan Blue (TB) before sample acquisition. P values were calculated using a t test between the indicated conditions.
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