English: Light-induced membrane recruitment of Raf1 resulted in prolonged retainment of nuclear ERK2-GFP. NIH3T3 cells were triply transfected with CIBN-CaaX, CRY2PHR-mCherry-Raf1, and ERK2-GFP. An epi-fluorescence microscope (Leica DMI6000B) equipped with an oil-immersion 100× objective was used for image acquisition. The blue light (∼472 nm) filtered from a mercury lamp was used as the stimulation light, whose power density was adjusted to ∼1 W/cm2 at the sample plane. Pulsed blue light (100-ms pulse duration, 30-s interval) was used for stimulation and imaging. After each pulse of blue light stimulation, a green light pulse was applied to acquire the fluorescence image of CRY2PHR-mCherry-Raf1. The same field of view is shown in Fig. 3A.