scholarly journals Two-Photon Excitation of Azobenzene Photoswitches for Synthetic Optogenetics

2020 ◽  
Vol 10 (3) ◽  
pp. 805 ◽  
Author(s):  
Shai Kellner ◽  
Shai Berlin
Keyword(s):  
Visible Light ◽  
Light Illumination ◽  
Optical Technique ◽  
Tissue Penetration ◽  
Cellular Targets ◽  
Two Photon ◽  
Photon Excitation ◽  
Two Photon Excitation

Synthetic optogenetics is an emerging optical technique that enables users to photocontrol molecules, proteins, and cells in vitro and in vivo. This is achieved by use of synthetic chromophores—denoted photoswitches—that undergo light-dependent changes (e.g., isomerization), which are meticulously designed to interact with unique cellular targets, notably proteins. Following light illumination, the changes adopted by photoswitches are harnessed to affect the function of nearby proteins. In most instances, photoswitches absorb visible light, wavelengths of poor tissue penetration, and excessive scatter. These shortcomings impede their use in vivo. To overcome these challenges, photoswitches of red-shifted absorbance have been developed. Notably, this shift in absorbance also increases their compatibility with two-photon excitation (2PE) methods. Here, we provide an overview of recent efforts devoted towards optimizing azobenzene-based photoswitches for 2PE and their current applications.

Quantitative determination of localized tissue oxygen concentration in vivo by two-photon excitation phosphorescence lifetime measurements

2004 ◽  
Vol 97 (5) ◽  
pp. 1962-1969 ◽  
Author(s):  
Egbert G. Mik ◽  
Ton G. van Leeuwen ◽  
Nicolaas J. Raat ◽  
Can Ince
Keyword(s):  
Rat Kidney ◽  
Excitation Wavelength ◽  
Lifetime Measurements ◽  
Phosphorescence Lifetime ◽  
Oxygen Determination ◽  
Two Photon ◽  
Photon Excitation ◽  
Two Photon Excitation

This study describes the use of two-photon excitation phosphorescence lifetime measurements for quantitative oxygen determination in vivo. Doubling the excitation wavelength of Pd-porphyrin from visible light to the infrared allows for deeper tissue penetration and a more precise and confined selection of the excitation volume due to the nonlinear two-photon effect. By using a focused laser beam from a 1,064-nm Q-switched laser, providing 10-ns pulses of 10 mJ, albumin-bound Pd-porphyrin was effectively excited and oxygen-dependent decay of phosphorescence was observed. In vitro calibration of phosphorescence lifetime vs. oxygen tension was performed. The obtained calibration constants were kq = 356 Torr−1·s−1 (quenching constant) and τ0 = 550 μs (lifetime at zero-oxygen conditions) at 37°C. The phosphorescence intensity showed a squared dependency to the excitation intensity, typical for two-photon excitation. In vivo demonstration of two-photon excitation phosphorescence lifetime measurements is shown by step-wise Po2 measurements through the cortex of rat kidney. It is concluded that quantitative oxygen measurements can be made, both in vitro and in vivo , using two-photon excitation oxygen-dependent quenching of phosphorescence. The use of two-photon excitation has the potential to lead to new applications of the phosphorescence lifetime technique, e.g., noninvasive oxygen scanning in tissue at high spatial resolution. To our knowledge, this is the first report in which two-photon excitation is used in the setting of oxygen-dependent quenching of phosphorescence lifetime measurements.

Two-Photon Excitation of Potentiometric Probes Enables Optical Recording of Action Potentials From Mammalian Nerve Terminals In Situ

2008 ◽  
Vol 99 (3) ◽  
pp. 1545-1553 ◽  
Author(s):  
Jonathan A. N. Fisher ◽  
Jonathan R. Barchi ◽  
Cristin G. Welle ◽  
Gi-Ho Kim ◽  
Paul Kosterin ◽  
...  
Keyword(s):  
Action Potentials ◽  
Optical Recording ◽  
Photon Absorption ◽  
Nerve Terminals ◽  
Electrode Arrays ◽  
Two Photon ◽  
Photon Excitation ◽  
Mammalian Nerve ◽  
Two Photon Excitation

We report the first optical recordings of action potentials, in single trials, from one or a few (∼1–2 μm) mammalian nerve terminals in an intact in vitro preparation, the mouse neurohypophysis. The measurements used two-photon excitation along the “blue” edge of the two-photon absorption spectrum of di-3-ANEPPDHQ (a fluorescent voltage-sensitive naphthyl styryl-pyridinium dye), and epifluorescence detection, a configuration that is critical for noninvasive recording of electrical activity from intact brains. Single-trial recordings of action potentials exhibited signal-to-noise ratios of ∼5:1 and fractional fluorescence changes of up to ∼10%. This method, by virtue of its optical sectioning capability, deep tissue penetration, and efficient epifluorescence detection, offers clear advantages over linear, as well as other nonlinear optical techniques used to monitor voltage changes in localized neuronal regions, and provides an alternative to invasive electrode arrays for studying neuronal systems in vivo.

In vivo label-free two-photon excitation autofluorescence microscopy of microvasculature using a 520 nm femtosecond fiber laser

2020 ◽  
Vol 45 (10) ◽  
pp. 2704
Author(s):  
Ting Wu ◽  
Jiuling Liao ◽  
Jia Yu ◽  
Yufeng Gao ◽  
Hui Li ◽  
...  
Keyword(s):  
Fiber Laser ◽  
Label Free ◽  
Two Photon ◽  
Photon Excitation ◽  
Femtosecond Fiber Laser ◽  
Two Photon Excitation

Intravital bone imaging by two-photon excitation microscopy to identify osteocytic osteolysis in vivo

Bone ◽  
2015 ◽  
Vol 74 ◽  
pp. 134-139 ◽  
Author(s):  
Hiroshige Sano ◽  
Junichi Kikuta ◽  
Masayuki Furuya ◽  
Naoki Kondo ◽  
Naoto Endo ◽  
...  
Keyword(s):  
Bone Imaging ◽  
Two Photon ◽  
Photon Excitation ◽  
Osteocytic Osteolysis ◽  
Two Photon Excitation

scholarly journals A novel method for observing proteins in vivo using a small fluorescent label and multiphoton imaging

2005 ◽  
Vol 390 (3) ◽  
pp. 787-790 ◽  
Author(s):  
Stanley W. Botchway ◽  
Ignasi Barba ◽  
Randolf Jordan ◽  
Rebecca Harmston ◽  
Peter M. Haggie ◽  
...  
Keyword(s):  
Fluorescence Detection ◽  
Fluorescence Emission ◽  
Yeast Cells ◽  
Fluorescent Label ◽  
Multiphoton Imaging ◽  
Two Photon ◽  
Photon Excitation ◽  
Novel Method ◽  
Two Photon Excitation

A novel method for the fluorescence detection of proteins in cells is described in the present study. Proteins are labelled by the selective biosynthetic incorporation of 5-hydroxytryptophan and the label is detected via selective two-photon excitation of the hydroxyindole and detection of its fluorescence emission at 340 nm. The method is demonstrated in this paper with images of a labelled protein in yeast cells.

scholarly journals In vivo live-cell imaging in plant tissues by two-photon excitation microscopy

2014 ◽  
Vol 26 (1) ◽  
pp. 25-30
Author(s):  
Yoko Mizuta ◽  
Daisuke Kurihara ◽  
Tetsuya Higashiyama
Keyword(s):  
Live Cell Imaging ◽  
Cell Imaging ◽  
Live Cell ◽  
Plant Tissues ◽  
Two Photon ◽  
Photon Excitation ◽  
Two Photon Excitation

scholarly journals Interactions of elastic and rigid vesicles with human skin in vitro: electron microscopy and two-photon excitation microscopy

1999 ◽  
Vol 1461 (1) ◽  
pp. 155-173 ◽  
Author(s):  
Benedicte A.I. van den Bergh ◽  
Jurrien Vroom ◽  
Hans Gerritsen ◽  
Hans E. Junginger ◽  
Joke A. Bouwstra
Keyword(s):  
Electron Microscopy ◽  
Human Skin ◽  
Two Photon ◽  
Photon Excitation ◽  
Two Photon Excitation
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