Third-generation GFP biosensors for real-time readout of pH and redox potential in living cells

Stephen James Remington; George T. Hanson; Mark Canon; Robert Aggeler; Devin Oglesbe; Roderick A. Capaldi; Roger Y. Tsien

doi:10.1117/12.530285

14 June 2004 Third-generation GFP biosensors for real-time readout of pH and redox potential in living cells

Stephen James Remington, George T. Hanson, Mark Canon, Robert Aggeler, Devin Oglesbe, Roderick A. Capaldi, Roger Y. Tsien

Author Affiliations +

Proceedings Volume 5329, Genetically Engineered and Optical Probes for Biomedical Applications II; (2004) https://doi.org/10.1117/12.530285
Event: Biomedical Optics 2004, 2004, San Jose, CA, United States

Abstract

Green fluorescent protein variants have been developed that report real-time change in pH and redox potential in living cells. The variants involve cysteine substitutions near the chromophore, which greatly alter the sensitivity of the protein to changes in its environment. Measurements can be made on single living cells in the fluorescence microscope or in cell suspension with an ordinary fluorimeter. The indicators are ratiometric by emission and/or excitation, which means that measurements at two different wavelengths are sufficient to determine both the quantity being measured and the indicator GFP concentration. The photophysics of a novel blue/green dual emission GFP variant will be presented. The design principles, crystal structures and ultrafast spectroscopic analysis of probe response will be discussed in terms of atomic models involving excited state proton transfer. Some applications in living cells will be presented.

Citation Download Citation

Stephen James Remington, George T. Hanson, Mark Canon, Robert Aggeler, Devin Oglesbe, Roderick A. Capaldi, and Roger Y. Tsien "Third-generation GFP biosensors for real-time readout of pH and redox potential in living cells", Proc. SPIE 5329, Genetically Engineered and Optical Probes for Biomedical Applications II, (14 June 2004); https://doi.org/10.1117/12.530285

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