High Temporal Resolution Study of Phosphorylation Events in HEK Cells Using a Micromixer Microfluidic Device

We developed a microfluidic mixing device with fast resolution times to conduct studies on protein phosphorylation in human embryonic kidney cells (HEK) upon epidermal growth factor (EGF) stimulation. The microfluidic device, capable of changing the cells’ local environment in under 100 ms, comprised two mixing segments: a first to ensure cell stimulation, and a second to quench the reaction and to lyse the cells. Different time points of stimulation were tested (0, 0.5, 1.0 and 5.0 s). Using mass spectrometry we quantified changes in protein phosphorylation upon EGF stimulation. For instance, the EGF receptor and downstream signalling proteins showed enhanced phosphorylation already at very early time points of stimulation.