scholarly journals The DIVER Microscope for Imaging in Scattering Media

2019 ◽  
Vol 2 (2) ◽  
pp. 53 ◽  
Author(s):  
Alexander Dvornikov ◽  
Leonel Malacrida ◽  
Enrico Gratton
Keyword(s):  
Imaging System ◽  
Detection System ◽  
Spectral Response ◽  
Biological Tissues ◽  
Scattering Media ◽  
Entrance Angle ◽  
Detection Scheme ◽  
Two Photon ◽  
Two Photon Fluorescence ◽  
Photon Fluorescence

We describe an advanced DIVER (Deep Imaging Via Emission Recovery) detection system for two-photon fluorescence microscopy that allows imaging in multiple scattering media, including biological tissues, up to a depth of a few mm with micron resolution. This detection system is more sensitive to low level light signals than conventional epi-detection used in two-photon fluorescence microscopes. The DIVER detector efficiently collects scattered emission photons from a wide area of turbid samples at almost any entrance angle in a 2π spherical angle. Using an epi-detection scheme only photons coming from a relatively small area of a sample and at narrow acceptance angle can be detected. The transmission geometry of the DIVER imaging system makes it exceptionally suitable for Second and Third Harmonic Generation (SHG, THG) signal detection. It also has in-depth fluorescence lifetime imaging (FLIM) capability. Using special optical filters with sin-cos spectral response, hyperspectral analysis of images acquired in-depth in scattering media can be performed. The system was successfully employed in imaging of various biological tissues. The DIVER detector can be plugged into a standard microscope stage and used as an external detector with upright commercial two-photon microscopes.

scholarly journals Simultaneous Two-Photon Fluorescence Microscopy of NADH and FAD Using Pixel-to-Pixel Wavelength-Switching

2021 ◽  
Vol 9 ◽  
Author(s):  
Yifan Qin ◽  
Yuanqin Xia
Keyword(s):  
Biological Tissues ◽  
Wavelength Switching ◽  
Excitation Scheme ◽  
Excitation Efficiency ◽  
Two Photon ◽  
Electro Optic ◽  
Two Photon Fluorescence ◽  
Photon Fluorescence ◽  
Bleed Through ◽  
Electro Optic Modulator

Two-photon fluorescence (TPF) microscopy of intrinsic fluorophores provides physiological and pathological information from biological tissues. Reduced nicotinamide adenine dinucleotide (NADH) and flavin adenine dinucleotide (FAD) are two endogenous fluorescent coenzymes existing on the intracellular scale. Autofluorescence images of NADH and FAD have been applied to noninvasively record changes during metabolism, according to their distributions and concentrations. However, the widely used sequential (non-simultaneous) excitation scheme results in artifacts caused by sample motion or laser power fluctuation. The single-wavelength illumination scheme suffers from low excitation efficiency and spectral bleed-through. In this paper, we demonstrate a new imaging system simultaneously capturing autofluorescence images from NADH and FAD, with high excitation efficiency and negligible spectral bleed-through. Two temporally multiplexed and spatially overlapped excitation beams were achieved with fast-switching light paths based on an electro-optic modulator. The switching beams were centered at 750 and 860 nm, enabling independent excitations of NADH and FAD. Autofluorescence images of NADH and FAD were acquired at the wavelength ranges of 415–455 nm and 500–550 nm, respectively. The electro-optic modulator was synchronized with the pixel clock from the microscope, achieving pixel-to-pixel wavelength-switching. The capability of the system was demonstrated by performing TPF imaging of freshly excised mouse colon tissues. The microenvironment of the colon wall was depicted by the distributions of colonocytes, goblet cells, and crypts of Lieberkühn, and the relative concentrations of NADH and FAD were estimated. The experimental results show that the system can effectively perform simultaneous imaging of NADH and FAD, and is considered a promising tool for investigations into metabolism-associated processes and diseases.

Multi-Photon Fluorescence Spectroum of Common Nucleic Acid Probes

2000 ◽  
Vol 6 (S2) ◽  
pp. 820-821
Author(s):  
P. C. Cheng ◽  
B. L. Lin ◽  
F. J. Kao ◽  
C. K. Sun ◽  
I. Johnson
Keyword(s):  
Fluorescence Microscopy ◽  
Fluorescent Probes ◽  
Fluorescence Spectra ◽  
Near Infrared ◽  
Single Photon ◽  
Spectral Response ◽  
Two Photon ◽  
Photon Excitation ◽  
Two Photon Fluorescence ◽  
Photon Fluorescence

Fluorescent probes are commonly used in biological fluorescence microscopy for tracking specific structures and sub-cellular compartments, and for indicating cellular ionic conditions. Recent development in multi-photon fluorescence microscopy has greatly expanded the usage of fluorescent probes in biomedical research. Considering its non-linear nature, two-photon excitation may generate very different fluorescence spectral response in the sample when compared with single photon excitation. It is thus necessary to measure the two-photon spectra of various fluorescent probes, so that two-photon fluorescence microscopy may be operated effectively and the images properly interpreted. This report represents the first installment of a continued effort in characterizing the multi-photon fluorescence spectra of commonly used bio-probes.Two-photon fluorescence spectra excited with near infrared at 780nm were obtained with a SpectraPro-500 spectrophotometer (Acton Research) equipped with a TE-cooled PMT and coupled to a Spectra-Physics Tsunami Ti-sapphire laser pumped by a Coherent Verdi solid-state laser operated at 85MHz, l00fs pulse.

scholarly journals Voxx: a PC-based, near real-time volume rendering system for biological microscopy

2002 ◽  
Vol 282 (1) ◽  
pp. C213-C218 ◽  
Author(s):  
Jeffrey L. Clendenon ◽  
Carrie L. Phillips ◽  
Ruben M. Sandoval ◽  
Shiaofen Fang ◽  
Kenneth W. Dunn
Keyword(s):  
Real Time ◽  
Imaging System ◽  
Low Cost ◽  
Three Dimensional ◽  
Image Data ◽  
Complex Structures ◽  
Two Photon ◽  
Two Photon Fluorescence ◽  
Photon Fluorescence ◽  
Biological Image

Confocal and two-photon fluorescence microscopy have advanced the exploration of complex, three-dimensional biological structures at submicron resolution. We have developed a voxel-based three-dimensional (3-D) imaging program (Voxx) capable of near real-time rendering that runs on inexpensive personal computers. This low-cost interactive 3-D imaging system provides a powerful tool for analyzing complex structures in cells and tissues and encourages a more thorough exploration of complex biological image data.

Lysosomal targeted NIR photosensitizer for photodynamic therapy and two-photon fluorescent imaging

2021 ◽  
Author(s):  
Ruiyuan Liu ◽  
Yuping Zhou ◽  
Di Zhang ◽  
Genghan He ◽  
Chuang Liu ◽  
...  
Keyword(s):  
Photodynamic Therapy ◽  
Fluorescence Imaging ◽  
Near Infrared ◽  
Fluorescent Imaging ◽  
High Fidelity ◽  
Design And Synthesis ◽  
Two Photon ◽  
Two Photon Fluorescence ◽  
Photon Fluorescence

Design and synthesis of near-infrared (NIR) emissive fluorophore for imaging of organelle and photodynamic therapy has received enormous attention. Hence, NIR emissive fluorophore of high-fidelity lysosome targeting, two-photon fluorescence imaging,...

Sign in / Sign up

Export Citation Format

Share Document