Figure 3. Fluorescence from an isolated ArcLight' GFP domain and eGFP show significant fluctuation noise.
(A) Fluorescence from a single isolated GFP domain from ArcLight' with no attached voltage sensor. The fluorescence displays fluctuations between different levels and does not appear to be Poisson-distributed. The trace was filtered at 100 Hz. (B) Background fluorescence from a region of an image with no GFP molecule displays no significant autocorrelation at any non-zero lag. This suggests that most noise in this regime is due to white processes such as shot noise. (C) The fluorescence from a single isolated GFP domain of ArcLight' displays significant autocorrelation. This suggests that the fluorescence moves between distinct levels corresponding to different conformational, chemical, or electronic states of the GFP domain. Blue bars in B and C represent approximate 95% confidence intervals for a Gaussian-distributed white noise process. No filtering was applied to the data used for the autocorrelations. (D) Single molecules of wild-type eGFP also display fluctuation noise that appears qualitatively similar to that of ArcLight' GFP domains, suggesting that this noise may be present in many GFP derivatives. The trace was filtered at 100 Hz. ADU is analog-to-digital units, the output unit of the EMCCD camera.