Automatic segmentation of lysosomes and analysis of intracellular pH with Radachlorin photosensitizer and FLIM.

Autor: Belashov AV; Ioffe Institute, Russian Academy of Sciences, 26, Polytekhnicheskaya, St.Petersburg, 194021, Russia., Zhikhoreva AA; Ioffe Institute, Russian Academy of Sciences, 26, Polytekhnicheskaya, St.Petersburg, 194021, Russia., Salova AV; Institute of Cytology, Russian Academy of Sciences, Tikhoretsky Pr., 4, St. Petersburg, 194064, Russia., Belyaeva TN; Institute of Cytology, Russian Academy of Sciences, Tikhoretsky Pr., 4, St. Petersburg, 194064, Russia., Litvinov IK; Institute of Cytology, Russian Academy of Sciences, Tikhoretsky Pr., 4, St. Petersburg, 194064, Russia., Kornilova ES; Institute of Cytology, Russian Academy of Sciences, Tikhoretsky Pr., 4, St. Petersburg, 194064, Russia., Semenova IV; Ioffe Institute, Russian Academy of Sciences, 26, Polytekhnicheskaya, St.Petersburg, 194021, Russia. Electronic address: irina.semenova@mail.ioffe.ru., Vasyutinskii OS; Ioffe Institute, Russian Academy of Sciences, 26, Polytekhnicheskaya, St.Petersburg, 194021, Russia.
Jazyk: angličtina
Zdroj: Biochemical and biophysical research communications [Biochem Biophys Res Commun] 2024 May 28; Vol. 710, pp. 149835. Date of Electronic Publication: 2024 Mar 29.
DOI: 10.1016/j.bbrc.2024.149835
Abstrakt: We report application of the fluorescence lifetime imaging microscopy (FLIM) for analysis of distributions of intracellular acidity using a chlorin-e6 based photosensitizer Radachlorin. An almost two-fold increase of the photosensitizer fluorescence lifetime in alkaline microenvironments as compared to acidic ones allowed for clear distinguishing between acidic and alkaline intracellular structures. Clusterization of a phasor plot calculated from fits of the FLIM raw data by two Gaussian distributions provided accurate automatic segmentation of lysosomes featuring acidic contents. The approach was validated in colocalization experiments with LysoTracker fluorescence in living cells of four established lines. The dependence of photosensitizer fluorescence lifetime on microenvironment acidity allowed for estimation of pH inside the cells, except for the nuclei, where photosensitizer does not penetrate. The developed method is promising for combined application of the photosensitizer for both photodynamic treatment and diagnostics.
Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
(Copyright © 2024 Elsevier Inc. All rights reserved.)
Databáze: MEDLINE
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