Stage incubator

The microfluidic cell migration experiment was per formed following the similar method described previously (Nandagopal et al., 2011) . The fluidic channel was coated with fibronectin (BD Biosciences, MA) for 1 h at room temperature and blocked with BSA for another hour before the experiment. For each experiment, cells were loaded into the microfluidic device from the wells and allowed to settle in the fibronectin coated channel for ~5 min. The device was maintained at 37 °C and 5% CO 2 using a stage incubator (PECON, Germany). Transfectants migration was imaged using a 10× objective with a CSU X1M 5000 spinning disc confocal microscope (Carl Zeiss Canada). Cell migration in the applied gradient fields was recorded by time lapse microscopy at 1 frame/min for 15 100 min using a CCD camera (Q32511, QIMAGING, Canada). A 63× objective was used to image CCR7 expression and internalization. A 488 nm solid state laser was used for imaging EGFP expression of the CCR7 transfectants and a 561 nm solid state laser was used for imaging RFP expression of the LifeAct transfectants. DIC images were taken in parallel. The image acquisition was controlled by ZEN2012 (Zeiss, Germany).