scholarly journals Volumetric two-photon imaging in live cells and embryos via axially gradient excitation

2019 ◽  
Author(s):  
Yufeng Gao ◽  
Xianyuan Xia ◽  
Jia Yu ◽  
Tingai Chen ◽  
Zhili Xu ◽  
...  
Keyword(s):  
Live Cells ◽  
Deep Penetration ◽  
Layer By Layer ◽  
Photon Beams ◽  
Volumetric Imaging ◽  
Localization Accuracy ◽  
Two Photon Microscopy ◽  
Two Photon ◽  
Subcellular Resolution ◽  
Imaging Speed

AbstractTwo-photon microscopy(TPM) that features subcellular resolution, intrinsic optical sectioning ability, and deep penetration in sample is a powerful tool of bioimaging. However, the process of layer-by-layer scanning to form a 3D image inherently limits the volumetric imaging speed and significantly increases the phototoxicity. Here we develop a gradient TPM technique that enables rapid volumetric imaging by only acquiring two 2D images. By sequentially exciting the specimen with two axially elongated two-photon beams with complementary gradient intensities, the axial positions of fluorophores can be decoded from the intensity ratio of the paired images. We achieve an axial localization accuracy of 0.728 ± 0.657 μm, which is sufficient for rapid 3D subcellular imaging. We demonstrate the flexibility of the gradient TPM on a variety of sparsely labelled samples, including bead phantoms, mouse brain tissues, live macrophages and live nematode embryos. The results show that, compared with conventional TPM, the 3D imaging speed increases 6 folds while the photobleaching and photodamage are extremely reduced.

Intravital multiphoton microscopy of dynamic renal processes

2005 ◽  
Vol 288 (6) ◽  
pp. F1084-F1089 ◽  
Author(s):  
Bruce A. Molitoris ◽  
Ruben M. Sandoval
Keyword(s):  
Proximal Tubule ◽  
Multiphoton Microscopy ◽  
Field Of View ◽  
Glomerular Permeability ◽  
Structural Alterations ◽  
Two Photon Microscopy ◽  
Two Photon ◽  
Dynamic Events ◽  
Subcellular Resolution ◽  
Functioning Kidney

Recent advances in microscopy and optics, computer sciences, and the available fluorophores used to label molecules of interest have empowered investigators to utilize intravital two-photon microscopy to study the dynamic events within the functioning kidney. This emerging technique enables investigators to follow functional and structural alterations with subcellular resolution within the same field of view over seconds to weeks. This approach invigorates the validity of data and facilitates analysis and interpretation as trends are more readily determined when one is more closely monitoring indicative physiological parameters. Therefore, in this review we emphasize how specific approaches will enable studies into glomerular permeability, proximal tubule endocytosis, and microvascular function within the kidney. We attempt to show how visual data can be quantified, thus allowing enhanced understanding of the process under study. Finally, emphasis is given to the possible future opportunities of this technology and its present limitations.

scholarly journals Poor man's two photon imaging: Scanning Laser Upconversion Microscopy

2017 ◽  
Author(s):  
Cecilia Sorbello ◽  
Roberto Etchenique
Keyword(s):  
Femtosecond Lasers ◽  
Laser Diodes ◽  
Scanning Laser ◽  
Deep Penetration ◽  
Multiphoton Absorption ◽  
Two Photon Microscopy ◽  
Two Photon ◽  
Instantaneous Power ◽  
Optical Images ◽  
Photon Imaging

ABSTRACTTwo photon microscopy is one of the most powerful techniques for optical images. Its deep penetration, low scattering and mainly its sectioning power allows exquisite control on all three axes of 3D samples, both for imaging and actuation. Unfortunately, this technique involves very expensive femtosecond lasers. Upconverting Nanoparticles display multiphoton absorption at low instantaneous power, which can be achieved by means of cheap laser diodes. Scanning Laser Upconversion Microscopy offers the deep penetration and low scattering of NIR excitation together with the sectioning power of multiphoton techniques for under US$ 1000.

scholarly journals Two-photon microscopy at >500 volumes/second

2020 ◽  
Author(s):  
Yu-Hsuan Tsai ◽  
Chih-Wei Liu ◽  
Wei-Kuan Lin ◽  
Chien-Sheng Wang ◽  
Chi-Huan Chiang ◽  
...  
Keyword(s):  
Data Rate ◽  
Two Photon Microscopy ◽  
Two Photon ◽  
Acoustic Lens ◽  
Axial Scanning ◽  
Imaging Speed

We demonstrate a multi-focal multi-photon volumetric microscopy via combination of 32-beam parallel lateral-scanning, a 70-kHz axial-scanning acoustic lens, and a 32-channel photodetector, enabling unprecedented data rate (2-10 GHz) and >500-volumes/second imaging speed over ~200×200×200-μm3.

Axial gradient excitation accelerates volumetric imaging of two-photon microscopy

2022 ◽  
Author(s):  
Yufeng Gao ◽  
Xianyuan Xia ◽  
Lina Liu ◽  
Chen Ting Ai ◽  
Ting Wu ◽  
...  
Keyword(s):  
Volumetric Imaging ◽  
Two Photon Microscopy ◽  
Two Photon ◽  
Axial Gradient

scholarly journals Molecular Tattooing of Live Cells and Animals: Spatial and Time Specific Effects of Azidoblebbistatin by Two-Photon Microscopy

2014 ◽  
Vol 106 (2) ◽  
pp. 177a
Author(s):  
Miklos Kepiro ◽  
Boglarka Varkuti ◽  
Gyorgy Hegyi ◽  
Miklos S.Z. Kellermayer ◽  
Malnasi-Csizmadia Andras
Keyword(s):  
Live Cells ◽  
Two Photon Microscopy ◽  
Two Photon ◽  
Specific Effects ◽  
Time Specific

scholarly journals Deep Penetration Microscopic Imaging with Non-Diffracting Airy Beams

Membranes ◽  
2021 ◽  
Vol 11 (6) ◽  
pp. 391
Author(s):  
Yong Guo ◽  
Yangrui Huang ◽  
Jin Li ◽  
Luwei Wang ◽  
Zhigang Yang ◽  
...  
Keyword(s):  
3D Structure ◽  
Kidney Tissue ◽  
Mouse Kidney ◽  
Deep Penetration ◽  
Imaging Method ◽  
Layer By Layer ◽  
Microscopic Imaging ◽  
Volumetric Imaging ◽  
Airy Beam ◽  
Volume Imaging

We report a deep penetration microscopic imaging method with a non-diffracting Airy beam. The direct mapping of volume imaging in free space shows that the axial imaging range of the Airy beam is approximately 4 times that of the traditional Gaussian beam along the axial direction while maintaining a narrow lateral width. Benefiting from its non-diffracting property, the microscopic imaging with Airy beam illumination can acquire image structures through turbid medium and capture a volumetric image in a single frame. We demonstrate the penetration ability of the Airy microscopic imaging through a strongly scattering environment with 633 nm and 780 nm lasers. The performances of the volumetric imaging method were evaluated using HeLa cells and isolated mouse kidney tissue. The thick sample was scanned layer by layer in the Gaussian mode, however, in the Airy mode, the three-dimensional (3D) structure information was projected onto a two-dimensional (2D) image, which vastly increased the volume imaging speed. To show the characteristics of the Airy microscope, we performed dynamic volumetric imaging on the isolated mouse kidney tissue with two-photon.

scholarly journals A FRET Based Two-Photon Fluorescent Probe for Visualizing Mitochondrial Thiols of Living Cells and Tissues

Sensors ◽  
2020 ◽  
Vol 20 (6) ◽  
pp. 1746 ◽  
Author(s):  
Zhengkun Liu ◽  
Qianqian Wang ◽  
Hao Wang ◽  
Wenting Su ◽  
Shouliang Dong
Keyword(s):  
High Selectivity ◽  
Limit Of Detection ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Fast Response ◽  
Live Cells ◽  
Biological Processes ◽  
Two Photon Microscopy ◽  
Two Photon ◽  
Main Component

Glutathione (GSH) is the main component of the mitochondrial thiol pool and plays key roles in the biological processes. Many evidences have suggested that cysteine and homocysteine also exist in mitochondria and are interrelated with GSH in biological systems. The fluctuation of the levels of mitochondrial thiols has been linked to many diseases and cells’ dysfunction. Therefore, the monitoring of mitochondrial thiol status is of great significance for clinical studies. We report here a novel fluorescence resonance energy transfer based two-photon probe MT-1 for mitochondrial thiols detection. MT-1 was constructed by integrating the naphthalimide moiety (donor) and rhodamine B (accepter and targeting group) through a newly designed linker. MT-1 shows a fast response, high selectivity, and sensitivity to thiols, as well as a low limit of detection. The two-photon property of MT-1 allows the direct visualization of thiols in live cells and tissues by two-photon microscopy. MT-1 can serve as an effective tool to unravel the diverse biological functions of mitochondrial thiols in living systems.

Two‐Photon Probes for Zn 2+ Ions with Various Dissociation Constants. Detection of Zn 2+ Ions in Live Cells and Tissues by Two‐Photon Microscopy

2011 ◽  
Vol 6 (5) ◽  
pp. 1234-1240 ◽  
Author(s):  
Isravel Antony Danish ◽  
Chang Su Lim ◽  
Yu Shun Tian ◽  
Ji Hee Han ◽  
Min Young Kang ◽  
...  
Keyword(s):  
Dissociation Constants ◽  
Live Cells ◽  
Two Photon Microscopy ◽  
Two Photon

scholarly journals Intravital imaging of adriamycin-induced renal pathology using two-photon microscopy and optical coherence tomography

2018 ◽  
Vol 11 (05) ◽  
pp. 1850030 ◽  
Author(s):  
Hengchang Guo ◽  
Hsing-Wen Wang ◽  
Qinggong Tang ◽  
Erik Anderson ◽  
Reuben Falola ◽  
...  
Keyword(s):  
Optical Coherence Tomography ◽  
Real Time ◽  
Flow Dynamics ◽  
Renal Pathology ◽  
Glomerular Sclerosis ◽  
Deep Penetration ◽  
Optical Coherence ◽  
Two Photon Microscopy ◽  
Two Photon ◽  
Tubular Flow

Adriamycin (doxorubicin), a common cancer chemotherapeutic drug, can be used to induce a model of chronic progressive glomerular disease in rodents. In our studies, we evaluated renal changes in a rat model after Adriamycin injection using two-photon microscopy (TPM), optical coherence tomography (OCT) and Doppler OCT (DOCT). Taking advantage of deep penetration and fast scanning speed for three-dimensional (3D) label-free imaging, OCT/DOCT system was able to reveal glomerular and tubular pathology noninvasively and in real time. By imaging renal pathology following the infusion of fluorophore-labeled dextrans of different molecular weights, TPM can provide direct views of glomerular and tubular flow dynamics with the onset and progression of renal disease. Specifically, glomerular permeability and filtration, proximal and distal tubular flow dynamics can be revealed. 6–8 weeks after injection of Adriamycin, TPM and OCT/DOCT imaging revealed glomerular sclerosis, compromised flow across the glomerular wall, tubular atrophy, tubular dilation, and variable intra-tubular flow dynamics. Our results indicate that TPM and OCT/DOCT provide real-time imaging of renal pathology in vivo that has not been previously available using conventional microscopic procedures.

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