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Article

FLIM-Based Intracellular and Extracellular pH Measurements Using Genetically Encoded pH Sensor

1
Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry of the RAS, 117997 Moscow, Russia
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A. N. Nesmeyanov Institute of Organoelement Compounds of the RAS, 119991 Moscow, Russia
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Prokhorov General Physics Institute of the RAS, 119991 Moscow, Russia
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Department of Biology, M.V. Lomonosov Moscow State University, 119234 Moscow, Russia
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A.V. Shubnikov Institute of Crystallography of Federal Scientific Research Centre “Crystallography and Photonics” of Russian Academy of Sciences, 119991 Moscow, Russia
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Sirius University of Science and Technology, 354340 Sochi, Russia
*
Author to whom correspondence should be addressed.
Biosensors 2021, 11(9), 340; https://doi.org/10.3390/bios11090340
Received: 9 August 2021 / Revised: 7 September 2021 / Accepted: 10 September 2021 / Published: 15 September 2021
(This article belongs to the Special Issue Advances of Genetically Encoded Biosensors)
The determination of pH in live cells and tissues is of high importance in physiology and cell biology. In this report, we outline the process of the creation of SypHerExtra, a genetically encoded fluorescent sensor that is capable of measuring extracellular media pH in a mildly alkaline range. SypHerExtra is a protein created by fusing the previously described pH sensor SypHer3s with the neurexin transmembrane domain that targets its expression to the cytoplasmic membrane. We showed that with excitation at 445 nm, the fluorescence lifetime of both SypHer3s and SypHerExtra strongly depend on pH. Using FLIM microscopy in live eukaryotic cells, we demonstrated that SypHerExtra can be successfully used to determine extracellular pH, while SypHer3s can be applied to measure intracellular pH. Thus, these two sensors are suitable for quantitative measurements using the FLIM method, to determine intracellular and extracellular pH in a range from pH 7.5 to 9.5 in different biological systems. View Full-Text
Keywords: GFP; SypHer3s; SypHerExtra; fluorescent proteins; pH; pH sensor; intracellular pH; extracellular pH; FLIM GFP; SypHer3s; SypHerExtra; fluorescent proteins; pH; pH sensor; intracellular pH; extracellular pH; FLIM
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MDPI and ACS Style

Goryashchenko, A.S.; Pakhomov, A.A.; Ryabova, A.V.; Romanishkin, I.D.; Maksimov, E.G.; Orsa, A.N.; Serova, O.V.; Mozhaev, A.A.; Maksimova, M.A.; Martynov, V.I.; Petrenko, A.G.; Deyev, I.E. FLIM-Based Intracellular and Extracellular pH Measurements Using Genetically Encoded pH Sensor. Biosensors 2021, 11, 340. https://doi.org/10.3390/bios11090340

AMA Style

Goryashchenko AS, Pakhomov AA, Ryabova AV, Romanishkin ID, Maksimov EG, Orsa AN, Serova OV, Mozhaev AA, Maksimova MA, Martynov VI, Petrenko AG, Deyev IE. FLIM-Based Intracellular and Extracellular pH Measurements Using Genetically Encoded pH Sensor. Biosensors. 2021; 11(9):340. https://doi.org/10.3390/bios11090340

Chicago/Turabian Style

Goryashchenko, Alexander S., Alexey A. Pakhomov, Anastasia V. Ryabova, Igor D. Romanishkin, Eugene G. Maksimov, Alexander N. Orsa, Oxana V. Serova, Andrey A. Mozhaev, Margarita A. Maksimova, Vladimir I. Martynov, Alexander G. Petrenko, and Igor E. Deyev 2021. "FLIM-Based Intracellular and Extracellular pH Measurements Using Genetically Encoded pH Sensor" Biosensors 11, no. 9: 340. https://doi.org/10.3390/bios11090340

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