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Label-Free Multiphoton Microscopy: The Origin of Fluorophores and Capabilities for Analyzing Biochemical Processes

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Abstract

Multiphoton microscopy (MPM) is a method of molecular imaging and specifically of intravital imaging that is characterized by high spatial resolution in combination with a greater depth of penetration into the tissue. MPM is a multimodal method based on detection of nonlinear optical signals–multiphoton fluorescence and optical harmonics–and also allows imaging with the use of the parameters of fluorescence decay kinetics. This review describes and discusses photophysical processes within major reporter molecules used in MPM with endogenous contrasts and summarizes several modern experiments that illustrate the capabilities of label-free MPM for molecular imaging of biochemical processes in connective tissue and cells.

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Abbreviations

AF:

autofluorescence

AGEs:

advanced glycation endproducts

CARS:

coherent anti-Stokes Raman scattering

FAD:

flavin adenine dinucleotide

FLIM:

fluorescence lifetime imaging

FP:

flavoprotein

IR:

infrared

MI:

metabolic imaging

MPM:

multiphoton microscopy

NAD:

nicotinamide adenine dinucleotide

NADF/NADB:

free/bound form of NAD

NADP:

nicotinamide adenine dinucleotide phosphate

OP:

oxidative phosphorylation

ROS:

reactive oxygen species

SC:

stem cells

SHG:

second harmonic generation

TCSPC:

time-correlated single photon counting

THG:

third harmonic generation

TP:

two-photon

UV:

ultraviolet

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Authors and Affiliations

  1. Lomonosov Moscow State University, Faculty of Physics, 119991, Moscow, Russia

    E. A. Shirshin, B. P. Yakimov, V. V. Fadeev & A. V. Priezzhev

  2. Institute of Spectroscopy, Russian Academy of Sciences, 108840, Troitsk, Moscow, Russia

    E. A. Shirshin

  3. Center of Experimental and Applied Cutaneous Physiology, Department of Dermatology, Venerology and Allergology, Charité–Universitätsmedizin Berlin, corporate member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health, Charitéplatz 1, 10117, Berlin, Germany

    M. E. Darvin & J. Lademann

  4. N. N. Priorov National Medical Research Center of Traumatology and Orthopaedics, 127299, Moscow, Russia

    N. P. Omelyanenko & S. A. Rodionov

  5. Medical Scientific–Educational Center of Lomonosov Moscow State University, 119192, Moscow, Russia

    Y. I. Gurfinkel

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  8. V. V. Fadeev
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Russian Text © E. A. Shirshin, B. P. Yakimov, M. E. Darvin, N. P. Omelyanenko, S. A. Rodionov, Y. I. Gurfinkel, J. Lademann, V. V. Fadeev, A. V. Priezzhev, 2019, published in Uspekhi Biologicheskoi Khimii, 2019, Vol. 59, pp. 139–180.

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Shirshin, E.A., Yakimov, B.P., Darvin, M.E. et al. Label-Free Multiphoton Microscopy: The Origin of Fluorophores and Capabilities for Analyzing Biochemical Processes. Biochemistry Moscow 84 (Suppl 1), 69–88 (2019). https://doi.org/10.1134/S0006297919140050

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  • DOI: https://doi.org/10.1134/S0006297919140050

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