Threshold illumination for non-invasive imaging of cells and tissues

Fluorescent microscopy employs monochromatic light which can affect the cells being observed. We reported earlier that fibroblasts relax their contractile force in response to green light of typical intensity. Here we show that such effects are independent of extracellular matrix and type of cell. In addition, we establish a threshold light that invokes minimal effect on cells. We cultured fibroblasts on soft 2D elastic hydrogels embedded with fluorescent beads to trace substrate deformation. The beads move towards cell center when cells contract, but they move away when cells relax. We use relaxation/contraction ratio,{lambda} r, as a measure of cell response to light. The cells were exposed to green (wavelength,{lambda} = 545-580 nm) and red ({lambda} = 635-650 nm) light with a range of intensities. We find red light with intensity less than ~ 57 W/m2 results in{lambda} r = 1, i.e., cells maintain force homeostasis. Higher intensities and smaller wavelengths result in widespread force-relaxation in cells with{lambda} r > 1. We suggest the use of{lambda} > 650 nm light with low intensity (I [≤] 57 W/m2) for time-lapse imaging of cells and tissues in order to avoid light-induced artifacts in experimental observations.