Two-Color Two-Photon Fluorescence Laser Scanning Microscopy

Quentmeier, Stefan; Denicke, Stefan; Gericke, Karl‐Heinz

doi:10.1007/s10895-009-0503-x

Cited by 32 publications

(27 citation statements)

References 26 publications

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“…The main benefit of 3-PF is the excitation of molecules that absorb in the UV (e.g. the amino acid tryptophane at 266 nm), which allows label-free imaging of intrinsic protein fluorescence [168]. Other authors have used 3-PF for label-free imaging of neurotransmitters such as serotonin and dopamine, which has implications for neuronal TE [157,169].…”

Section: Three-photon-based Nonlinear Optical Microscopy Techniquesmentioning

confidence: 99%

Taking a deep look: modern microscopy technologies to optimize the design and functionality of biocompatible scaffolds for tissue engineering in regenerative medicine

Vielreicher

Schürmann

Detsch

et al. 2013

J. R. Soc. Interface.

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This review focuses on modern nonlinear optical microscopy (NLOM) methods that are increasingly being used in the field of tissue engineering (TE) to image tissue non-invasively and without labelling in depths unreached by conventional microscopy techniques. With NLOM techniques, biomaterial matrices, cultured cells and their produced extracellular matrix may be visualized with high resolution. After introducing classical imaging methodologies such as µCT, MRI, optical coherence tomography, electron microscopy and conventional microscopy two-photon fluorescence (2-PF) and second harmonic generation (SHG) imaging are described in detail (principle, power, limitations) together with their most widely used TE applications. Besides our own cell encapsulation, cell printing and collagen scaffolding systems and their NLOM imaging the most current research articles will be reviewed. These cover imaging of autofluorescence and fluorescence-labelled tissue and biomaterial structures, SHG-based quantitative morphometry of collagen I and other proteins, imaging of vascularization and online monitoring techniques in TE. Finally, some insight is given into state-of-the-art three-photon-based imaging methods (e.g. coherent anti-Stokes Raman scattering, third harmonic generation). This review provides an overview of the powerful and constantly evolving field of multiphoton microscopy, which is a powerful and indispensable tool for the development of artificial tissues in regenerative medicine and which is likely to gain importance also as a means for general diagnostic medical imaging.

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Section: Three-photon-based Nonlinear Optical Microscopy Techniquesmentioning

confidence: 99%

Taking a deep look: modern microscopy technologies to optimize the design and functionality of biocompatible scaffolds for tissue engineering in regenerative medicine

Vielreicher

Schürmann

Detsch

et al. 2013

J. R. Soc. Interface.

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“…Lim and Saloma have predicted that error caused by spherical aberration is reduced with ND-2PE versus D-2PE [31]. Quentmeier et al [32,33] demonstrated UVfluorescence lifetime measurements using ND-2PE in the visible spectrum, and Robinson et al [34] demonstrated fluorescence lifetime imaging using ND-2PE combining visible and NIR wavelengths. Recently, Mahou et al demonstrated that ND-2PE can be used for simultaneous excitation of four different fluorescent proteins in mouse cerebral cortex [35].…”

Section: Introductionmentioning

confidence: 99%

Non-degenerate 2-photon excitation in scattering medium for fluorescence microscopy

Yang¹,

Abashin²,

Saisan³

et al. 2016

Opt. Express

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Non-degenerate 2-photon excitation (ND-2PE) of a fluorophore with two laser beams of different photon energies offers an independent degree of freedom in tuning of the photon flux for each beam. This feature takes advantage of the infrared wavelengths used in degenerate 3-photon excitation (D-3PE) microscopy to achieve increased penetration depths, while preserving a relatively high 2-photon excitation cross section in comparison to that of D-3PE. Here, using spatially and temporally aligned Ti:Sapphire laser and optical parametric oscillator beams operating at near infrared (NIR) and short-wavelength infrared (SWIR) optical frequencies, we employ ND-2PE and provide a practical demonstration that a constant fluorophore emission intensity is achievable deeper into a scattering medium using ND-2PE as compared to the commonly used degenerate 2-photon excitation (D-2PE). References and links1. K. Svoboda and R. Yasuda, "Principles of two-photon excitation microscopy and its applications to neuroscience," Neuron 50(6), 823-839 (2006). 2. J. N. Kerr and W. Denk, "Imaging in vivo: watching the brain in action," Nat. Rev. Neurosci. 9(3), 195-205 (2008). 3. W. Denk and K. Svoboda, "Photon upmanship: why multiphoton imaging is more than a gimmick," Neuron 18(3), 351-357 (1997). 4. F. Helmchen and W. Denk, "Deep tissue two-photon microscopy," Nat. Methods 2(12), 932-940 (2005 11. D. Kobat, N. G. Horton, and C. Xu, "In vivo two-photon microscopy to 1.6-mm depth in mouse cortex," J. Biomed. Opt. 16(10), 106014 (2011). 12. R. Kawakami, K. Sawada, A. Sato, T. Hibi, Y. Kozawa, S. Sato, H. Yokoyama, and T. Nemoto, "Visualizing hippocampal neurons with in vivo two-photon microscopy using a 1030 nm picosecond pulse laser," Sci. Rep. 3, 1014 (2013). 13. P. Theer and W. Denk, "On the fundamental imaging-depth limit in two-photon microscopy," J. Opt. Soc. Am.A adaptive optical recovery of optimal resolution over large volumes," Nat. Methods 11(6), 625-628 (2014).

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“…The power of triple two-photon excitation was demonstrated to intravitally visualize three chromophores and second harmonic generation (SHG) in fly embryos 44 , and seven fluorophores in distinct cellular and tissue compartment in murine lymph nodes 38 . The triple two-photon excitation scheme is based on a wavelength mixing technique 45,46 , which implies temporal synchronization of pulse trains from two lasers with the same repetition rate as well as spatial overlap of their foci volumes (Figure 1a). The SHG can be detected from a tissue with ordered structure, i.e.…”

mentioning

confidence: 99%

New dimensions in intravital multi-photon imaging of immune reactions

Niesner¹

2017

J Immunological Sci

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In the last two decades intravital multi-photon imaging has become a central tool to investigate cellular and molecular dynamics of immune reactions in vivo. Currently, challenges in exploiting the full power of this technology include limitations on the number of simultaneously detectable parameters as well as in expanding the acquisition in time and space. Here we discuss technological advancements developed in order to overcome these challenges and focus on the example of germinal center reactions as multi-parametric immunological processes evolving over a time course of days and weeks.

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Two-Color Two-Photon Fluorescence Laser Scanning Microscopy

Cited by 32 publications

References 26 publications

Taking a deep look: modern microscopy technologies to optimize the design and functionality of biocompatible scaffolds for tissue engineering in regenerative medicine

Taking a deep look: modern microscopy technologies to optimize the design and functionality of biocompatible scaffolds for tissue engineering in regenerative medicine

Non-degenerate 2-photon excitation in scattering medium for fluorescence microscopy

New dimensions in intravital multi-photon imaging of immune reactions

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