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Fig. 5. ECAR increases in response to increasing endogenous and exogenous EGF. (A) Cells lacking EGF expression were grown in transwells and placed in a cytosensor. Once a baseline ECAR was established, the cells were challenged with an addition of exogenous EGF, 7 nM ({circ}) or 0.1 nM (). The reported ECAR is normalized to the baseline signal that was set to 0 for clarity. (B) Cells were induced to secrete ligand at a steady state rate of 0 ({circ}), 45 (), 400 ({square}), 700 ({blacksquare}), 1300 ({triangleup}), 3600 ({blacktriangleup}) moleculesxcell-1xminute-1 using tetracycline and a ligand-release inhibitor. The cells were placed on the cytosensor and once a baseline was established, they were challenged with 40, 20, 5, 2.5, 1.2, 0.6, 0.3, 0.15, 0.08 ng/ml EGF. The maximal change in ECAR, normalized to baseline, was recorded as a function of the challenge concentration. The error bars represent the s.e.m. of three experiments in duplicate.