Label-Free Non-linear Multimodal Optical Microscopy—Basics, Development, and Applications

Nirmal Mazumder, Naveen K. Balla, Guan Yu Zhuo, Yury V. Kistenev, Rajesh Kumar, Fu Jen Kao, Sophie Brasselet, Viktor V. Nikolaev, Natalya A. Krivova

Research output: Contribution to journalReview article

Abstract

Non-linear optical (NLO) microscopy has proven to be a powerful tool especially for tissue imaging with sub-cellular resolution, high penetration depth, endogenous contrast specificity, pinhole-less optical sectioning capability. In this review, we discuss label-free non-linear optical microscopes including the two-photon fluorescence (TPF), fluorescence lifetime imaging microscopy (FLIM), polarization-resolved second harmonic generation (SHG) and coherent anti-Stokes Raman scattering (CARS) techniques with various samples. The non-linear signals are generated from collagen in tissue (SHG), amylopectin from starch granules (SHG), sarcomere structure of fresh muscle (SHG), elastin in skin (TPF), nicotinamide adenine dinucleotide (NADH) in cells (TPF), and lipid droplets in cells (CARS). Again, the non-linear signals are very specific to the molecular structure of the sample and its relative orientation to the polarization of the incident light. Thus, polarization-resolved non-linear optical microscopy provides high image contrast and quantitative estimate of sample orientation. An overview of the advancements on polarization-resolved SHG microscopy including Stokes vector based polarimetry, circular dichroism, and susceptibility are also presented in this review article. The working principles and corresponding implements of above-mentioned microscopy techniques are elucidated. The potential of time-resolved TPF lifetime imaging microscopy (TP-FLIM) is explored by imaging endogenous fluorescence of NAD(P)H, a key coenzyme in cellular metabolic processes. We also discuss single laser source time-resolved multimodal CARS-FLIM microscopy using time-correlated single-photon counting (TCSPC) in combination with continuum generation from photonic crystal fiber (PCF). Using examples, we demonstrate that the multimodal NLO microscopy is a powerful tool to assess the molecular specificity with high resolution.

Original languageEnglish
Article number170
JournalFrontiers in Physics
Volume7
DOIs
Publication statusPublished - 31-10-2019

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Microscopy
Labels
Fluorescence
Microscopic examination
microscopy
Photons
Harmonic generation
Optical Imaging
Second Harmonic Generation
fluorescence
Imaging techniques
Fluorescence Lifetime
Stokes
Raman Spectrum Analysis
Imaging
Raman Spectra
NAD
Polarization
Optical microscopy
Raman scattering

All Science Journal Classification (ASJC) codes

  • Biophysics
  • Materials Science (miscellaneous)
  • Mathematical Physics
  • Physics and Astronomy(all)
  • Physical and Theoretical Chemistry

Cite this

Mazumder, N., Balla, N. K., Zhuo, G. Y., Kistenev, Y. V., Kumar, R., Kao, F. J., ... Krivova, N. A. (2019). Label-Free Non-linear Multimodal Optical Microscopy—Basics, Development, and Applications. Frontiers in Physics, 7, [170]. https://doi.org/10.3389/fphy.2019.00170
Mazumder, Nirmal ; Balla, Naveen K. ; Zhuo, Guan Yu ; Kistenev, Yury V. ; Kumar, Rajesh ; Kao, Fu Jen ; Brasselet, Sophie ; Nikolaev, Viktor V. ; Krivova, Natalya A. / Label-Free Non-linear Multimodal Optical Microscopy—Basics, Development, and Applications. In: Frontiers in Physics. 2019 ; Vol. 7.
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Mazumder, N, Balla, NK, Zhuo, GY, Kistenev, YV, Kumar, R, Kao, FJ, Brasselet, S, Nikolaev, VV & Krivova, NA 2019, 'Label-Free Non-linear Multimodal Optical Microscopy—Basics, Development, and Applications', Frontiers in Physics, vol. 7, 170. https://doi.org/10.3389/fphy.2019.00170

Label-Free Non-linear Multimodal Optical Microscopy—Basics, Development, and Applications. / Mazumder, Nirmal; Balla, Naveen K.; Zhuo, Guan Yu; Kistenev, Yury V.; Kumar, Rajesh; Kao, Fu Jen; Brasselet, Sophie; Nikolaev, Viktor V.; Krivova, Natalya A.

In: Frontiers in Physics, Vol. 7, 170, 31.10.2019.

Research output: Contribution to journalReview article

TY - JOUR

T1 - Label-Free Non-linear Multimodal Optical Microscopy—Basics, Development, and Applications

AU - Mazumder, Nirmal

AU - Balla, Naveen K.

AU - Zhuo, Guan Yu

AU - Kistenev, Yury V.

AU - Kumar, Rajesh

AU - Kao, Fu Jen

AU - Brasselet, Sophie

AU - Nikolaev, Viktor V.

AU - Krivova, Natalya A.

PY - 2019/10/31

Y1 - 2019/10/31

N2 - Non-linear optical (NLO) microscopy has proven to be a powerful tool especially for tissue imaging with sub-cellular resolution, high penetration depth, endogenous contrast specificity, pinhole-less optical sectioning capability. In this review, we discuss label-free non-linear optical microscopes including the two-photon fluorescence (TPF), fluorescence lifetime imaging microscopy (FLIM), polarization-resolved second harmonic generation (SHG) and coherent anti-Stokes Raman scattering (CARS) techniques with various samples. The non-linear signals are generated from collagen in tissue (SHG), amylopectin from starch granules (SHG), sarcomere structure of fresh muscle (SHG), elastin in skin (TPF), nicotinamide adenine dinucleotide (NADH) in cells (TPF), and lipid droplets in cells (CARS). Again, the non-linear signals are very specific to the molecular structure of the sample and its relative orientation to the polarization of the incident light. Thus, polarization-resolved non-linear optical microscopy provides high image contrast and quantitative estimate of sample orientation. An overview of the advancements on polarization-resolved SHG microscopy including Stokes vector based polarimetry, circular dichroism, and susceptibility are also presented in this review article. The working principles and corresponding implements of above-mentioned microscopy techniques are elucidated. The potential of time-resolved TPF lifetime imaging microscopy (TP-FLIM) is explored by imaging endogenous fluorescence of NAD(P)H, a key coenzyme in cellular metabolic processes. We also discuss single laser source time-resolved multimodal CARS-FLIM microscopy using time-correlated single-photon counting (TCSPC) in combination with continuum generation from photonic crystal fiber (PCF). Using examples, we demonstrate that the multimodal NLO microscopy is a powerful tool to assess the molecular specificity with high resolution.

AB - Non-linear optical (NLO) microscopy has proven to be a powerful tool especially for tissue imaging with sub-cellular resolution, high penetration depth, endogenous contrast specificity, pinhole-less optical sectioning capability. In this review, we discuss label-free non-linear optical microscopes including the two-photon fluorescence (TPF), fluorescence lifetime imaging microscopy (FLIM), polarization-resolved second harmonic generation (SHG) and coherent anti-Stokes Raman scattering (CARS) techniques with various samples. The non-linear signals are generated from collagen in tissue (SHG), amylopectin from starch granules (SHG), sarcomere structure of fresh muscle (SHG), elastin in skin (TPF), nicotinamide adenine dinucleotide (NADH) in cells (TPF), and lipid droplets in cells (CARS). Again, the non-linear signals are very specific to the molecular structure of the sample and its relative orientation to the polarization of the incident light. Thus, polarization-resolved non-linear optical microscopy provides high image contrast and quantitative estimate of sample orientation. An overview of the advancements on polarization-resolved SHG microscopy including Stokes vector based polarimetry, circular dichroism, and susceptibility are also presented in this review article. The working principles and corresponding implements of above-mentioned microscopy techniques are elucidated. The potential of time-resolved TPF lifetime imaging microscopy (TP-FLIM) is explored by imaging endogenous fluorescence of NAD(P)H, a key coenzyme in cellular metabolic processes. We also discuss single laser source time-resolved multimodal CARS-FLIM microscopy using time-correlated single-photon counting (TCSPC) in combination with continuum generation from photonic crystal fiber (PCF). Using examples, we demonstrate that the multimodal NLO microscopy is a powerful tool to assess the molecular specificity with high resolution.

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