Fig. 5.

GFP lifetime accuracy in live cell imaging under different laser excitation power. (a) Histograms of GFP lifetime distribution narrow with increasing excitation power. (b) The standard deviation of GFP lifetime is inversely proportional to the square root of excitation power. (c) GFP lifetime image of HeLa cells at 200 μW excitation power. (d) GFP lifetime image of HeLa cells at 30 μW excitation power. GFP fluorescence lifetime was measured at 2.25 ± 0.16 ns at 200 μW excitation power and 2.3 ± 0.3 ns at 30 μW excitation power. Image size is 400-by-400 pixels with 0.3 μm pixel size. The image set was acquired at full 44,000 pixel/s speed. Lifetimes were fitted from data smoothed by a 2-by-2 pixel wide Gaussian window. The intensity image was processed at the original resolution without smoothing.