Figure 3. VEGFA₁₆₅b is a weaker activator of AKT in primary HRMECs compared to VEGFA₁₆₅a. A: Activation of AKT was measured with in situ immunofluorescence with a phosphospecific antibody for Ser473-AKT at five time points after the addition of 20 ng/ml (1,050 pM) VEGFA₁₆₅a or VEGFA₁₆₅b. Bar shows standard deviation. (t test, ***p<0.001 relative to untreated control, n=4 biologic replicates.) B: Dose–response curves for activation of AKT. The median effective dose (ED₅₀) value for AKT activation by VEGFA₁₆₅a was 53 pM compared to 126 pM for VEGFA₁₆₅b. Bars indicate the 95% confidence interval for data fit to the four-parameter log-logistic function. n=4 biologic replicates per dose.