The human epidermal growth factor receptor (HER) system is an intricately regulated system that plays critical roles in development and tumorigenesis. Here, we apply integrated experimentation and modeling to analyze HER receptor activation in a panel of cell lines expressing endogenous levels of EGFR/HER1 and different levels of HER2. A mathematical model that includes the fundamental processes involved in receptor activation and trafficking was used to fit the experimental data, and values of the independent parameters for active receptor dimer formation affinities, trafficking rates and relative phosphorylation levels were estimated. Obtained parameter values quantitatively support the existing ideas on the effect of HER2 on EGFR dynamics, and enable us to predict the abundances of various phosphorylated receptor dimers in the cell lines.