Optically Modulated Photoswitchable Fluorescent Proteins Yield Improved Biological Imaging Sensitivity.
Optically Modulated Photoswitchable Fluorescent Proteins Yield Improved Biological Imaging Sensitivity.
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DOI：
10.1021/jacs.5b07871
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发表时间：
2015-10-14
影响因子：
15
通讯作者：
Dickson RM
中科院分区：
文献类型：
--
作者：
Chen YC;Jablonski AE;Issaeva I;Bourassa D;Hsiang JC;Fahrni CJ;Dickson RM
Photoswitchable fluorescent proteins (PS-FPs) open grand new opportunities in biological imaging. Through optical manipulation of FP emission, we demonstrate that dual-laser modulated synchronously amplified fluorescence image recovery (DM-SAFIRe) improves signal contrast in high background through unambiguous demodulation and is linear in relative fluorophore abundance at different points in the cell. The unique bright-to-dark state interconversion rates of each PS-FP not only enables discrimination of different, yet spectrally indistinguishable FPs, but also allows signal rejection of diffusing relative to bound forms of the same PS-FP, rsFastLime. Adding to the sensitivity gains realized from rejecting non-modulatable background, the selective signal recovery of immobilized vs. diffusing intracellular rsFastLime suggests that DM-SAFIRe can detect weak protein-protein interactions that are normally obscured by large fractions of unbound fluorescent proteins.
