News Imagerie cellulaire - Cellular imaging
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Segmentation and Shape Analysis of Macrophages Using Anglegram Analysis

Segmentation and Shape Analysis of Macrophages Using Anglegram Analysis | News Imagerie cellulaire - Cellular imaging | Scoop.it

José Alonso Solís-Lemus, Brian Stramer, Greg Slabaugh and Constantino Carlos Reyes-Aldasoro


Cell migration is crucial in many processes of development and maintenance of multicellular organisms and it can also be related to disease, e.g., Cancer metastasis, when cells migrate to organs different to where they originate. A precise analysis of the cell shapes in biological studies could lead to insights about migration. However, in some cases, the interaction and overlap of cells can complicate the detection and interpretation of their shapes. This paper describes an algorithm to segment and analyse the shape of macrophages in fluorescent microscopy image sequences, and compares the segmentation of overlapping cells through different algorithms. A novel 2D matrix with multiscale angle variation, called the anglegram, based on the angles between points of the boundary of an object, is used for this purpose. The anglegram is used to find junctions of cells and applied in two different applications: (i) segmentation of overlapping cells and for non-overlapping cells; (ii) detection of the “corners” or pointy edges in the shapes. The functionalities of the anglegram were tested and validated with synthetic data and on fluorescently labelled macrophages observed on embryos of Drosophila melanogaster. The information that can be extracted from the anglegram shows a good promise for shape determination and analysis, whether this involves overlapping or non-overlapping objects.


J. Imaging 2018, 4(1), 2;

doi:10.3390/jimaging4010002


http://www.mdpi.com/2313-433X/4/1/2

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Wide-field multiphoton imaging through scattering media without correction

Wide-field multiphoton imaging through scattering media without correction | News Imagerie cellulaire - Cellular imaging | Scoop.it

Adrià Escobet-Montalbán, Roman Spesyvtsev, Mingzhou Chen, Wardiya Afshar Saber, Melissa Andrews, C. Simon Herrington, Michael Mazilu and Kishan Dholakia

 

Optical approaches to fluorescent, spectroscopic, and morphological imaging have made exceptional advances in the last decade. Super-resolution imaging and wide-field multiphoton imaging are now underpinning major advances across the biomedical sciences. While the advances have been startling, the key unmet challenge to date in all forms of optical imaging is to penetrate deeper. A number of schemes implement aberration correction or the use of complex photonics to address this need. In contrast, we approach this challenge by implementing a scheme that requires no a priori information about the medium nor its properties. Exploiting temporal focusing and single-pixel detection in our innovative scheme, we obtain wide-field two-photon images through various turbid media including a scattering phantom and tissue reaching a depth of up to seven scattering mean free path lengths. Our results show that it competes favorably with standard point-scanning two-photon imaging, with up to a fivefold improvement in signal-to-background ratio while showing significantly lower photobleaching.

 

Science Advances, 12 Oct 2018 : Vol. 4, no. 10, eaau1338
DOI: 10.1126/sciadv.aau1338

Open Access : http://advances.sciencemag.org/content/advances/4/10/eaau1338.full.pdf

 

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Invited Article: Label-free nerve imaging with a coherent anti-Stokes Raman scattering rigid endoscope using two optical fibers for laser delivery.

Invited Article: Label-free nerve imaging with a coherent anti-Stokes Raman scattering rigid endoscope using two optical fibers for laser delivery. | News Imagerie cellulaire - Cellular imaging | Scoop.it

Keigo Hirose, Shuichiro Fukushima, Taichi Furukawa, Hirohiko Niioka and Mamoru Hashimoto

 

A coherent anti-Stokes Raman scattering (CARS) rigid endoscope using two optical fibers to deliver excitation beams individually is developed. The use of two optical fibers allows the correction of longitudinal chromatic aberration and enhances the CARS signal by a factor of 2.59. The endoscope is used to image rat sciatic nerves with an imaging time of 10 s. Imaging of the rabbit prostatic fascia without sample slicing is also demonstrated, which reveals the potential for the application of the CARS endoscope to robot-assisted surgery.
 

APL Photonics, Volume 3, Issue 9 >10.1063/1.5031817

https://doi.org/10.1063/1.5031817

http://aip.scitation.org/doi/pdf/10.1063/1.5031817?class=pdf

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Visualizing protein synthesis in mice with in vivo labeling of deuterated amino acids using vibrational imaging

Visualizing protein synthesis in mice with in vivo labeling of deuterated amino acids using vibrational imaging | News Imagerie cellulaire - Cellular imaging | Scoop.it

Lingyan Shi, Yihui Shen and Wei Min

 

Proteins are one of the major components of biological systems, and understanding their metabolism is critical to study various biochemical processes in living systems. Despite extensive efforts to study protein metabolism such as autoradiography, mass spectrometry, and fluorescence microscopy, visualizing the spatial distribution of overall protein metabolism in mammals at subcellular resolution is still challenging. A recent study from our group reported imaging newly synthesized proteins in cultured mammalian cells, tissues, or even in mice using stimulated Raman scattering (SRS) microscopy coupled with metabolic labeling of deuterated amino acids (dAA). However, our previous method of dAA administration via drinking water, albeit convenient, is insufficient for in vivo studies. This is due to poor labeling efficiency and limited access to many important organs such as the brain, pancreas, or tumor. In this study, we have significantly improved and optimized the in vivo administration method by intra-carotid arterial injection of dAA in mice and obtained imaging contrast of protein metabolic activity in many more organs and tissues, such as cerebral and cerebellar cortex and hippocampal regions in the mouse brain. We also imaged newly formed proteins in the choroid plexus and pancreas at different time points, illustrating the metabolic dynamics of proteins in these important secretory organs. In addition, we visualized the metabolic heterogeneity of protein synthesis in colon tumor xenografts, which can be used to distinguish tumor and normal tissues. In summary, this combination of a new dAA administration technique and SRS imaging platform demonstrates an effective tool for the in vivo study of complex protein metabolism in mammals, in both physiological and pathological states.
 

APL Photonic, Volume 3, Issue 9 >10.1063/1.5028134

https://doi.org/10.1063/1.5028134

Open Access : http://aip.scitation.org/doi/pdf/10.1063/1.5028134?class=pdf

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Perspective: Coherent Raman scattering microscopy, the future is bright

Perspective: Coherent Raman scattering microscopy, the future is bright | News Imagerie cellulaire - Cellular imaging | Scoop.it

Chi Zhang and Ji-Xin Cheng

 

Chemical imaging offers critical information to understand the fundamentals in biology and to assist clinical diagnostics. Label-free chemical imaging piques a general interest since it avoids the use of bio-perturbing molecular labels and holds promises to characterize human tissue in vivo. Coherent Raman scattering (CRS), which utilizes lasers to excite the vibrations of molecules, renders new modalities to map chemicals in living samples without the need of labeling and provides significantly improved speed, resolution, and sensitivity compared to spontaneous Raman scattering. Although microscopy systems based on CRS have seen rapid development in the past two decades, remaining challenges, which emerge in diverse aspects, start to impede the continuous advancement of the field. In this perspective, we review the history of CRS microscopy, scrutinize the pros and cons of different modalities, and discuss the current challenges and possible future directions of the field. Infiltration of conceptual and technological ideals from other fields will promote CRS microscopy towards a versatile tool for basic science and medical research.

 

APL Photonics, Volume 3, Issue 9 

https://doi.org/10.1063/1.5040101

Open Access : https://aip.scitation.org/doi/pdf/10.1063/1.5040101?class=pdf

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High Spatiotemporal Resolution Imaging with Localized Plasmonic Structured Illumination Microscopy

High Spatiotemporal Resolution Imaging with Localized Plasmonic Structured Illumination Microscopy | News Imagerie cellulaire - Cellular imaging | Scoop.it
Anna Bezryadina, Junxiang Zhao, Yang Xia, Xiang Zhang and Zhaowei Liu
 

Localized plasmonic structured illumination microscopy (LPSIM) provides multicolor wide-field super-resolution imaging with low phototoxicity and high-speed capability. LPSIM utilizes a nanoscale plasmonic antenna array to provide a series of tunable illumination patterns beyond the traditional diffraction limit, allowing for enhanced resolving powers down to a few tens of nanometers. Here, we demonstrate wide-field LPSIM with 50 nm spatial resolution at video rate speed by imaging microtubule dynamics with low illumination power intensity. The design of the LPSIM system makes it suitable for imaging surface effects of cells and tissues with regular sample preparation protocols. LPSIM can be extended to much higher resolution, representing an excellent technology for live-cell imaging of protein dynamics and interactions.

 

ACS Nano, 2018, 12 (8), pp 8248–8254
DOI : 10.1021/acsnano.8b03477
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Aggregation-Induced Emission Luminogen with Near-Infrared-II Excitation and Near-Infrared-I Emission for Ultradeep Intravital Two-Photon Microscopy

Aggregation-Induced Emission Luminogen with Near-Infrared-II Excitation and Near-Infrared-I Emission for Ultradeep Intravital Two-Photon Microscopy | News Imagerie cellulaire - Cellular imaging | Scoop.it
Ji Qi, Chaowei Sun, Dongyu Li, Hequn Zhang, Wenbin Yu, Abudureheman Zebibula, Jacky W. Y. Lam, Wang Xi, Liang Zhu, Fuhong Cai, Peifa Wei, Chunlei Zhu, Ryan T. K. Kwok, Lina L. Streich, Robert Prevedel, Jun Qian and Ben Zhong Tang
 

Currently, a serious problem obstructing the large-scale clinical applications of fluorescence technique is the shallow penetration depth. Two-photon fluorescence microscopic imaging with excitation in the longer-wavelength near-infrared (NIR) region (>1100 nm) and emission in the NIR-I region (650–950 nm) is a good choice to realize deep-tissue and high-resolution imaging. Here, we report ultradeep two-photon fluorescence bioimaging with 1300 nm NIR-II excitation and NIR-I emission (peak ∼810 nm) based on a NIR aggregation-induced emission luminogen (AIEgen). The crab-shaped AIEgen possesses a planar core structure and several twisting phenyl/naphthyl rotators, affording both high fluorescence quantum yield and efficient two-photon activity. The organic AIE dots show high stability, good biocompatibility, and a large two-photon absorption cross section of 1.22 × 103 GM. Under 1300 nm NIR-II excitation, in vivo two-photon fluorescence microscopic imaging helps to reconstruct the 3D vasculature with a high spatial resolution of sub-3.5 μm beyond the white matter (>840 μm) and even to the hippocampus (>960 μm) and visualize small vessels of ∼5 μm as deep as 1065 μm in mouse brain, which is among the largest penetration depths and best spatial resolution of in vivo two-photon imaging. Rational comparison with the AIE dots manifests that two-photon imaging outperforms the one-photon mode for high-resolution deep imaging. This work will inspire more sight and insight into the development of efficient NIR fluorophores for deep-tissue biomedical imaging.

 

ACS Nano, 2018, 12 (8), pp 7936–7945
DOI : 10.1021/acsnano.8b02452
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Automated quantification of bioluminescence images

Automated quantification of bioluminescence images | News Imagerie cellulaire - Cellular imaging | Scoop.it

Alexander D. Klose & Neal Paragas

 

We developed a computer-aided analysis tool for quantitatively determining bioluminescent reporter distributions inside small animals. The core innovations are a body-fitting animal shuttle and a statistical mouse atlas, both of which are spatially aligned and scaled according to the animal’s weight, and hence provide data congruency across animals of varying size and pose. In conjunction with a multispectral bioluminescence tomography technique capitalizing on the spatial framework of the shuttle, the in vivo biodistribution of luminescent reporters can rapidly be calculated and, thus, enables operator-independent and computer-driven data analysis. We demonstrate its functionality by quantitatively monitoring a bacterial infection, where the bacterial organ burden was determined and validated with the established serial-plating method. In addition, the statistical mouse atlas was validated and compared to existing techniques providing an anatomical reference. The proposed data analysis tool promises to increase data throughput and data reproducibility and accelerate human disease modeling in mice.

 

Nature Communications volume 9, Article number: 4262 (2018)

https://doi.org/10.1038/s41467-018-06288-w

Open Access : https://www.nature.com/articles/s41467-018-06288-w.pdf

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Tissue-adhesive wirelessly powered optoelectronic device for metronomic photodynamic cancer therapy

Tissue-adhesive wirelessly powered optoelectronic device for metronomic photodynamic cancer therapy | News Imagerie cellulaire - Cellular imaging | Scoop.it
Kento Yamagishi, Izumi Kirino, Isao Takahashi, Hizuru Amano, Shinji Takeoka, Yuji Morimoto, Toshinori Fujie


Metronomic (that is, low-dose and long-term) photodynamic therapy (mPDT) for treating internal lesions requires the stable fixation of optical devices to internal tissue surfaces to enable continuous, local light delivery. Surgical suturing—the standard choice for device fixation—can be unsuitable in the presence of surrounding major nerves and blood vessels, as well as for organs or tissues that are fragile, change their shape or actively move. Here, we show that an implantable and wirelessly powered mPDT device consisting of near-field-communication-based light-emitting-diode chips and bioadhesive and stretchable polydopamine-modified poly(dimethylsiloxane) nanosheets can be stably fixed onto the inner surface of animal tissue. When implanted subcutaneously in mice with intradermally transplanted tumours, the device led to significant antitumour effects by irradiating for 10 d at approximately 1,000-fold lower intensity than conventional PDT approaches. The mPDT device might facilitate treatment strategies for hard-to-detect microtumours and deeply located lesions that are hard to reach with standard phototherapy.

 

Nature Biomedical Engineering (2018)

Doi : 10.1038/s41551-018-0261-7


Via Miguel Martín-Landrove
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Imaging the zebrafish, one cell at a time – Morgridge Institute for Research

Imaging the zebrafish, one cell at a time – Morgridge Institute for Research | News Imagerie cellulaire - Cellular imaging | Scoop.it

A new imaging project at the Morgridge Institute for Research might be the biology equivalent of a 19th century expressionist painting. Think Van Gogh’s “Starry Night,” a constellation of tiny lines of color combining into a powerful image.
Except the canvas of this research project will be a zebrafish, and the paint will be individual cells of a developing embryo.
Jan Huisken, Morgridge medical engineering investigator and visiting professor in the UW-Madison Department of Integrative Biology, is part of an ambitious project to develop a complete cellular blueprint of zebrafish development, from the first ball of cells to an adult fish. The project could have great benefit to regenerative biology, by precisely defining what role each individual cell plays in the full development of a complex organism.
The project is one of the 2018 winners of the “High-Risk, High-Reward Research Program” announced today (Oct. 2) by the National Institutes of Health (NIH). The project is one of 10 transformative research awards from the NIH Director’s Office that funds investigators whose research ideas “could potentially create or challenge existing paradigms.”
Three labs will contribute distinct expertise to the effort. Project lead David Traver, a biological science professor at the University of California-San Diego, develops unique tools to identify, colorfully visualize and track zebrafish stem cells. Zhirong Bao, a developmental biologist at the Sloan Kettering Institute, provides computational tools to track the movement and function of cells over time. And Huisken provides the imaging expertise through light sheet microscopy, which can non-invasively image living zebrafish embryos for as long as 48 hours.
Huisken says the project’s strongest suit is combining these three domains — labeling, imaging and tracking — to do something no single lab could do alone.
“We want to achieve something in the zebrafish that has only been achievable in much smaller organisms,” says Huisken. “We envision creating an atlas that scientists can look into and see how all of the cell lineages have taken shape.”
Today, the completion of developmental blueprints has been restricted to the model organism c. elegans, a relatively simple worm whose early development is scripted almost like a computation. But in more complex organisms like the zebrafish, there is far more variation from one individual to the next, making it harder to determine cellular fates.
“We envision creating an atlas that scientists can look into and see how all of the cell lineages have taken shape.”
This project will use laser marking systems to randomly assign a different color to each of hundreds of early-stage cells. Every cell will be a slightly different color than their neighbors, allowing researcher to track their migrations.
Interestingly, the daughter cells of each of these cells will carry on the same color as the parent. Huisken says this will result in a mosaic of color patterns across the fish. If a cluster of heart cells is a particular shade of green, they will be able to map those cells back to the original source.
The live imaging via light sheet microscopy can only be realistically done through the first few days of the embryo, Huisken says. At later stages of development to adulthood, the fish will be fixed, cleared and rapidly imaged with a different light sheet configuration.
Another unique contribution of the Huisken Lab will be a microscope nicknamed Flamingo. This iteration of a light sheet microscope is shrunk down to the size of a suitcase that can be shared with biology labs that have fragile specimens. The labs at UCSD and Sloan Kettering will be able to use Flamingo to perfect their methodologies, while the high-throughput imaging can concentrate at Morgridge.
Zebrafish area ideal model organisms for many reasons. They are translucent, grow rapidly and are highly prolific — a single couple can produce a thousand embryos a day. They provide a vivid window into how each organ is formed.

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Fluorescence Polarization Control for On–Off Switching of Single Molecules at Cryogenic Temperatures

Fluorescence Polarization Control for On–Off Switching of Single Molecules at Cryogenic Temperatures | News Imagerie cellulaire - Cellular imaging | Scoop.it
Christiaan N. Hulleman, Maximiliaan Huisman, Robert J. Moerland, David Grünwald, Sjoerd Stallinga, and Bernd Rieger

 

Light microscopy, allowing sub‐diffraction‐limited resolution, has been among the fastest developing techniques at the interface of biology, chemistry, and physics. Intriguingly no theoretical limit exists on how far the underlying measurement uncertainty can be lowered. In particular data fusion of large amounts of images can reduce the measurement error to match the resolution of structural methods like cryo‐electron microscopy. Fluorescence, although reliant on a reporter molecule and therefore not the first choice to obtain ultraresolution structures, brings highly specific labeling of molecules in a large assembly to the table and inherently allows the detection of multiple colors, which enables the interrogation of multiple molecular species at the same time in the same sample. Here, the problems to be solved in the coming years, with the aim of higher resolution, are discussed, and what polarization depletion of fluorescence at cryogenic temperatures can contribute for fluorescence imaging of biological samples, like whole cells, is described.

 

Small Methods 2018, 2, 1700323
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Absorption by water increases fluorescence image contrast of biological tissue in the shortwave infrared

Absorption by water increases fluorescence image contrast of biological tissue in the shortwave infrared | News Imagerie cellulaire - Cellular imaging | Scoop.it

Jessica A. Carr, Marianne Aellen, Daniel Franke, Peter T. C. So, Oliver T. Bruns, and Moungi G. Bawendi

 

Shortwave infrared (SWIR) fluorescence imaging is a tool for visualizing biological processes deep within tissue or living animals. Our study shows that the contrast in a SWIR fluorescence image is primarily mediated by the absorptivity of the tissue, and can therefore be tuned through deliberate selection of imaging wavelength. We show, for example, that, in 3D tissue phantoms and in brain vasculature in vivo in mice, imaging at SWIR wavelengths of the highest water absorptivity results in the greatest fluorescence contrast. We further demonstrate, in microscopy of ex vivo mouse liver tissue, that imaging at wavelengths of high tissue absorptivity can also increase imaging penetration depth, and use a theoretical contrast model to explain this effect.

 

PNAS September 11, 2018 115 (37) 9080-9085

https://doi.org/10.1073/pnas.1803210115

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Morphology of mitochondria in spatially restricted axons revealed by cryo-electron tomography

Morphology of mitochondria in spatially restricted axons revealed by cryo-electron tomography | News Imagerie cellulaire - Cellular imaging | Scoop.it

Tara D. Fischer, Pramod K. Dash, Jun Liu, M. Neal Waxham

 

Neurons project axons to local and distal sites and can display heterogeneous morphologies with limited physical dimensions that may influence the structure of large organelles such as mitochondria. Using cryo-electron tomography (cryo-ET), we characterized native environments within axons and presynaptic varicosities to examine whether spatial restrictions within these compartments influence the morphology of mitochondria. Segmented tomographic reconstructions revealed distinctive morphological characteristics of mitochondria residing at the narrowed boundary between presynaptic varicosities and axons with limited physical dimensions (approximately 80 nm), compared to mitochondria in nonspatially restricted environments. Furthermore, segmentation of the tomograms revealed discrete organizations between the inner and outer membranes, suggesting possible independent remodeling of each membrane in mitochondria at spatially restricted axonal/varicosity boundaries. Thus, cryo-ET of mitochondria within axonal subcompartments reveals that spatial restrictions do not obstruct mitochondria from residing within them, but limited available space can influence their gross morphology and the organization of the inner and outer membranes. These findings offer new perspectives on the influence of physical and spatial characteristics of cellular environments on mitochondrial morphology and highlight the potential for remarkable structural plasticity of mitochondria to adapt to spatial restrictions within axons.

 

PLoS Biol 16(9): e2006169.

DOI : 10.1371/journal.pbio.2006169

Open Access : https://journals.plos.org/plosbiology/article/file?id=10.1371/journal.pbio.2006169&type=printable

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Nanoscale imaging of the adhesion core including integrin β1 on intact living cells using scanning electron-assisted dielectric-impedance microscopy

Nanoscale imaging of the adhesion core including integrin β1 on intact living cells using scanning electron-assisted dielectric-impedance microscopy | News Imagerie cellulaire - Cellular imaging | Scoop.it

Tomoko Okada, Toshihiko Ogura

 

The integrins are a superfamily of transmembrane proteins composed of α and β subunit dimers involved in cell–cell and cell–extracellular matrix interactions. The largest integrin subgroup is integrin β1, which contributes to several malignant phenotypes. Recently, we have developed a novel imaging technology named scanning electron-assisted dielectric-impedance microscopy (SE-ADM), which visualizes untreated living mammalian cells in aqueous conditions with high contrast. Using the SE-ADM system, we observed 60-nm gold colloids with antibodies directly binding to the focal adhesion core containing integrin β1 on mammalian cancer cells without staining and fixation. The adhesion core contains three or four high-density regions of integrin β1 and connects to the actin filament. An adhesion core with high-density integrin β1 is suggested to contain 10–20 integrin dimers. Our SE-ADM system can also visualize various other membrane proteins in living cells in medium without staining and fixation.

 

PLoS ONE 13(9): e0204133.

DOI : 10.1371/journal.pone.0204133

Open Access : https://journals.plos.org/plosone/article/file?id=10.1371/journal.pone.0204133&type=printable

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Imaging beyond the super-resolution limits using ultrastructure expansion microscopy (UltraExM)

Imaging beyond the super-resolution limits using ultrastructure expansion microscopy (UltraExM) | News Imagerie cellulaire - Cellular imaging | Scoop.it

Davide Gambarotto, Fabian Zwettler, Marketa Cernohorska, Denis Fortun, Susanne Borgers, Jorn Heine, Jan-Gero Schloetel, Matthias Reuss, Michael Unser, Edward Boyden, Markus Sauer, Virginie Hamel, Paul Guichard

 

For decades, electron microscopy (EM) was the only method able to reveal the ultrastructure of cellular organelles and molecular complexes because of the diffraction limit of optical microscopy. In recent past, the emergence of super-resolution fluorescence microscopy enabled the visualization of cellular structures with so far unmatched spatial resolution approaching virtually molecular dimensions. Despite these technological advances, currently super-resolution microscopy does not permit the same resolution level as provided by electron microscopy, impeding the attribution of a protein to an ultrastructural element. Here, we report a novel method of near-native expansion microscopy (UltraExM), enabling the visualization of preserved ultrastructures of macromolecular assemblies with subdiffraction-resolution by standard optical microscopy. UltraExM revealed for the first time the ultrastructural localization of tubulin glutamylation in centrioles. Combined with super-resolution microscopy, UltraExM unveiled the centriolar chirality, an ultrastructural signature, which was only visualizable by electron microscopy.

 

bioRxiv preprint first posted online Apr. 25, 2018 not peer-reviewed

http://dx.doi.org/10.1101/308270

Open Access : https://www.biorxiv.org/content/biorxiv/early/2018/04/25/308270.full.pdf

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Invited Article: Comparison of hyperspectral coherent Raman scattering microscopies for biomedical applications.

Invited Article: Comparison of hyperspectral coherent Raman scattering microscopies for biomedical applications. | News Imagerie cellulaire - Cellular imaging | Scoop.it

T. Bocklitz, T. Meyer, M. Schmitt, I. Rimke, F. Hoffmann, F. von Eggeling, G. Ernst, O. Guntinas-Lichius and J. Popp

 

Raman scattering based imaging represents a very powerful optical tool for biomedical diagnostics. Different Raman signatures obtained by distinct tissue structures and disease induced changes provoke sophisticated analysis of the hyperspectral Raman datasets. While the analysis of linear Raman spectroscopic tissue data is quite established, the evaluation of hyperspectral nonlinear Raman data has not yet been evaluated in great detail. The two most common nonlinear Raman methods are CARS (coherent anti-Stokes Raman scattering) and SRS (stimulated Raman scattering) spectroscopy. Specifically the linear concentration dependence of SRS as compared to the quadratic dependence of CARS has fostered the application of SRS tissue imaging. Here, we applied spectral processing to hyperspectral SRS and CARS data for tissue characterization. We could demonstrate for the first time that similar cluster distributions can be obtained for multispectral CARS and SRS data but that clustering is based on different spectral features due to interference effects in CARS and the different concentration dependence of CARS and SRS. It is shown that a direct combination of CARS and SRS data does not improve the clustering results.

 

APL Photonics, Volume 3, Issue 9 >10.1063/1.5030159

https://doi.org/10.1063/1.5030159

Open Access : http://aip.scitation.org/doi/pdf/10.1063/1.5030159?class=pdf

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Tutorial: Coherent Raman light matter interaction processes

Tutorial: Coherent Raman light matter interaction processes | News Imagerie cellulaire - Cellular imaging | Scoop.it

Hervé  Rigneault and Pascal Berto

 

Coherent Raman scattering processes such as coherent anti-Stokes Raman scattering and stimulated Raman scattering are described in a tutorial way keeping simple physical pictures and simple derivations. The simplicity of the presentation keeps however most of the key features of these coherent and resonant processes and their intimate relation with spontaneous Raman scattering. This tutorial provides a digest of introduction to the fundamental physics at work, and it does not focus on the numerous technological implementations; rather, it provides the concepts and the physical tools to understand the extensive literature in this field. The presentation is made simple enough for under-graduate students, graduate students, and newcomers with various scientific backgrounds.

 

APL Photonics, Volume 3, Issue 9 > 10.1063/1.5030335

https://doi.org/10.1063/1.5030335

Open Access : https://aip.scitation.org/doi/pdf/10.1063/1.5030335?class=pdf

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Single-molecule analysis of endogenous β-actin mRNA trafficking reveals a mechanism for compartmentalized mRNA localization in axons

Single-molecule analysis of endogenous β-actin mRNA trafficking reveals a mechanism for compartmentalized mRNA localization in axons | News Imagerie cellulaire - Cellular imaging | Scoop.it

Benita Turner-Bridger, Maximillian Jakobs, Leila Muresan, Hovy Ho-Wai Wong, Kristian Franze, William A. Harris, and Christine E. Holt

 

De novo protein synthesis in neuronal axons plays important roles in neural circuit formation, maintenance, and disease. Key to the selectivity of axonal protein synthesis is whether an mRNA is present at the right place to be translated, but the mechanisms behind axonal mRNA localization remain poorly understood. In this work, we quantitatively analyze the link between axonal β-actin mRNA trafficking and its localization patterns. By developing a single-molecule approach to live-image β-actin mRNAs in axons, we explore the biophysical drivers behind β-actin mRNA motion and uncover a mechanism for generating increased density at the axon tip by differences in motor protein-driven transport speeds. These results provide mechanistic insight into the control of local translation through mRNA trafficking.

 

PNAS October 9, 2018 115 (41) E9697-E9706

https://doi.org/10.1073/pnas.1806189115 

Open Access : http://www.pnas.org/content/115/41/E9697.full.pdf

 

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Bright Near-Infrared Aggregation-Induced Emission Luminogens with Strong Two-Photon Absorption, Excellent Organelle Specificity, and Efficient Photodynamic Therapy Potential

Bright Near-Infrared Aggregation-Induced Emission Luminogens with Strong Two-Photon Absorption, Excellent Organelle Specificity, and Efficient Photodynamic Therapy Potential | News Imagerie cellulaire - Cellular imaging | Scoop.it
Zheng Zheng, Tianfu Zhang, Haixiang Liu, Yuncong Chen, Ryan T. K. Kwok, Chao Ma, Pengfei Zhang, Herman H. Y. Sung, Ian D. Williams, Jacky W. Y. Lam, Kam Sing Wong and Ben Zhong Tang
 

Far-red and near-infrared (NIR) fluorescent materials possessing the characteristics of strong two-photon absorption and aggregation-induced emission (AIE) as well as specific targeting capability are much-sought-after for bioimaging and therapeutic applications due to their deep penetration depth and high resolution. Herein, a series of dipolar far-red and NIR AIE luminogens with a strong push–pull effect are designed and synthesized. The obtained fluorophores display bright far-red and NIR solid-state fluorescence with a high quantum yield of up to 30%, large Stokes shifts of up to 244 nm, and large two-photon absorption cross-sections of up to 887 GM. A total of three neutral AIEgens show specific lipid droplet (LD)-targeting capability, while the one with cationic and lipophilic characteristics tends to target the mitochondria specifically. All of the molecules demonstrate good biocompatibility, high brightness, and superior photostability. They also serve as efficient two-photon fluorescence-imaging agents for the clear visualization of LDs or mitochondria in living cells and tissues with deep tissue penetration (up to 150 μm) and high contrast. These AIEgens can efficiently generate singlet oxygen upon light irradiation for the photodynamic ablation of cancer cells. All of these intriguing results prove that these far-red and NIR AIEgens are excellent candidates for the two-photon fluorescence imaging of LDs or mitochondria and organelle-targeting photodynamic cancer therapy.

 

ACS Nano, 2018, 12 (8), pp 8145–8159
DOI : 10.1021/acsnano.8b03138
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Tunable Aggregation-Induced Emission Nanoparticles by Varying Isolation Groups in Perylene Diimide Derivatives and Application in Three-Photon Fluorescence Bioimaging

Tunable Aggregation-Induced Emission Nanoparticles by Varying Isolation Groups in Perylene Diimide Derivatives and Application in Three-Photon Fluorescence Bioimaging | News Imagerie cellulaire - Cellular imaging | Scoop.it

Luyi Zong, Hequn Zhang, Yaqin Li, Yanbin Gong, Dongyu Li, Jiaqiang Wang, Zhe Wang, Yujun Xie, Mengmeng Han, Qian Peng, Xuefeng Li, Jinfeng Dong, Jun Qian, Qianqian Li and Zhen Li

 

The development of fluorogens with deep-red emission is one of the hottest topics of investigation in the field of bio/chemosensors and bioimaging. Herein, the tunable fluorescence of perylene diimide (PDI) derivatives was achieved by the incorporation of varied isolation groups linked on the PDI core. With the enlarged sizes of isolation groups, the conversion from aggregation caused quenching to aggregation-induced emission was obtained in their fluorescence variations from solutions to nanoparticles, as the result of the efficient inhibition of π–π stacking by the larger isolation groups. Accordingly, DCzPDI bearing 1,3-di(9H-carbazol-9-yl)benzene as the biggest isolation group exhibited the bright deep-red emission in the aggregated state with a quantum yield of 12.3%. Combined with the three-photon excited fluorescence microscopy (3PFM) technology, through-skull 3PFM imaging of mouse cerebral vasculature can be realized by DCzPDI nanoparticles with good biocompatibility, and the penetration depth can be as deep as 450 μm.

 

ACS Nano, 2018, 12 (9), pp 9532–9540
DOI : 10.1021/acsnano.8b05090

 

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Homotransfer FRET Reporters for Live Cell Imaging

Homotransfer FRET Reporters for Live Cell Imaging | News Imagerie cellulaire - Cellular imaging | Scoop.it

Nicole E. Snell, Vishnu P. Rao, Kendra M. Seckinger, Junyi Liang, Jenna Leser, Allison E. Mancini and M. A. Rizzo

 

Förster resonance energy transfer (FRET) between fluorophores of the same species was recognized in the early to mid-1900s, well before modern heterotransfer applications. Recently, homotransfer FRET principles have re-emerged in biosensors that incorporate genetically encoded fluorescent proteins. Homotransfer offers distinct advantages over the standard heterotransfer FRET method, some of which are related to the use of fluorescence polarization microscopy to quantify FRET between two fluorophores of identical color. These include enhanced signal-to-noise, greater compatibility with other optical sensors and modulators, and new design strategies based upon the clustering or dimerization of singly-labeled sensors. Here, we discuss the theoretical basis for measuring homotransfer using polarization microscopy, procedures for data collection and processing, and we review the existing genetically-encoded homotransfer biosensors.

 

Biosensors 2018, 8(4), 89

https://doi.org/10.3390/bios8040089

Open Access : https://www.mdpi.com/2079-6374/8/4/89/pdf

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Single‐Nanoparticle Cell Barcoding by Tunable FRET from Lanthanides to Quantum Dots

Single‐Nanoparticle Cell Barcoding by Tunable FRET from Lanthanides to Quantum Dots | News Imagerie cellulaire - Cellular imaging | Scoop.it

 

 

Chi Chen, Dr. Lijiao Ao, Yu‐Tang Wu, Vjona Cifliku, Dr. Marcelina Cardoso Dos Santos, Emmanuel Bourrier, Dr. Martina Delbianco, Prof. David Parker, Dr. Jurriaan M. Zwier, Dr. Liang Huang, Prof. Niko Hildebrandt
 

Fluorescence barcoding based on nanoparticles provides many advantages for multiparameter imaging. However, creating different concentration‐independent codes without mixing various nanoparticles and by using single‐wavelength excitation and emission for multiplexed cellular imaging is extremely challenging. Herein, we report the development of quantum dots (QDs) with two different SiO2 shell thicknesses (6 and 12 nm) that are coated with two different lanthanide complexes (Tb and Eu). FRET from the Tb or Eu donors to the QD acceptors resulted in four distinct photoluminescence (PL) decays, which were encoded by simple time‐gated (TG) PL intensity detection in three individual temporal detection windows. The well‐defined single‐nanoparticle codes were used for live cell imaging and a one‐measurement distinction of four different cells in a single field of view. This single‐color barcoding strategy opens new opportunities for multiplexed labeling and tracking of cells.

 

Angew.Chem. 2018, 130,13876–13881

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Strategies to Overcome Autofluorescence in Nanoprobe‐Driven In Vivo Fluorescence Imaging

Strategies to Overcome Autofluorescence in Nanoprobe‐Driven In Vivo Fluorescence Imaging | News Imagerie cellulaire - Cellular imaging | Scoop.it
Blanca del Rosal and Antonio Benayas

 

The development of fluorescent probes and optical detection systems in the near‐infrared (700–2000 nm) has boosted the interest in fluorescence bioimaging as an alternative to traditional medical imaging techniques. Fluorescence imaging can provide high‐resolution images at fast acquisition speeds, while removing the need for ionizing radiations or radioactive contrast agents and requiring relatively simple and cost‐effective equipment. The low absorption and scattering of near‐infrared radiation by biological tissues enables minimally invasive visualization of deeply embedded organs and structures. However, the infrared autofluorescence background generated by some biological components, as discussed here, can negatively affect the image contrast and complicate the visualization of the fluorescent probes used as contrast agents. A critical review on the different approaches for improving the signal‐to‐noise ratio in in vivo fluorescence imaging experiments through autofluorescence background removal is presented here.

 

Small Methods 2018, 2, 1800075
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Lighting Up MicroRNA in Living Cells by the Disassembly of Lock‐Like DNA‐Programmed UCNPs‐AuNPs through the Target Cycling Amplification Strategy

Lighting Up MicroRNA in Living Cells by the Disassembly of Lock‐Like DNA‐Programmed UCNPs‐AuNPs through the Target Cycling Amplification Strategy | News Imagerie cellulaire - Cellular imaging | Scoop.it
Keying Zhang, Shuting Song, Shan Huang, Lin Yang, Qianhao Min, Xingcai Wu, Feng Lu
 

Intracellular microRNAs imaging based on upconversion nanoprobes has great potential in cancer diagnostics and treatments. However, the relatively low detection sensitivity limits their application. Herein, a lock‐like DNA (LLD) generated by a hairpin DNA (H1) hybridizing with a bolt DNA (bDNA) sequence is designed, which is used to program upconversion nanoparticles (UCNPs, NaYF4@NaYF4:Yb, Er@NaYF4) and gold nanoparticles (AuNPs). The upconversion emission is quenched through luminescence resonance energy transfer (LRET). The multiple LLD can be repeatedly opened by one copy of target microRNA under the aid of fuel hairpin DNA strands (H2) to trigger disassembly of AuNPs from the UCNP, resulting in the lighting up of UCNPs with a high detection signal gain. This strategy is verified using microRNA‐21 as model. The expression level of microRNA‐21 in various cells lines can be sensitively measured in vitro, meanwhile cancer cells and normal cells can be easily and accurately distinguished by intracellular microRNA‐21 imaging via the nanoprobes. The detection limit is about 1000 times lower than that of the previously reported upconversion nanoprobes without signal amplification. This is the first time a nonenzymatic signal amplification method has been combined with UCNPs for imaging intracellular microRNAs, which has great potential for cancer diagnosis.

 

Small Volume14, Issue40, October 4, 2018,

1802292

https://doi.org/10.1002/smll.201802292

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Convolutional neural networks automate detection for tracking of submicron-scale particles in 2D and 3D

Convolutional neural networks automate detection for tracking of submicron-scale particles in 2D and 3D | News Imagerie cellulaire - Cellular imaging | Scoop.it

Jay M. Newby, Alison M. Schaefer, Phoebe T. Lee, M. Gregory Forest, and Samuel K. Lai

 

The increasing availability of powerful light microscopes capable of collecting terabytes of high-resolution 2D and 3D videos in a single day has created a great demand for automated image analysis tools. Tracking the movement of nanometer-scale particles (e.g., virus, proteins, and synthetic drug particles) is critical for understanding how pathogens breach mucosal barriers and for the design of new drug therapies. Our advancement is to use an artificial neural network that provides, first and foremost, substantially improved automation. Additionally, our method improves accuracy compared with current methods and reproducibility across users and laboratories.

 

PNAS September 4, 2018, 115 (36) 9026–9031

https://doi.org/10.1073/pnas.1804420115

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SYBR Gold dye enables preferential labelling of mitochondrial nucleoids and their time-lapse imaging by structured illumination microscopy

SYBR Gold dye enables preferential labelling of mitochondrial nucleoids and their time-lapse imaging by structured illumination microscopy | News Imagerie cellulaire - Cellular imaging | Scoop.it

Visnja Jevtic, Petra Kindle, Sergiy V. Avilov

 

Mitochondrial DNA molecules coated with proteins form compact particles called mitochondrial nucleoids. They are redistributed within mitochondrial network undergoing morphological changes. The straightforward technique to characterize nucleoids’ motions is fluorescence microscopy. Mitochondrial nucleoids are commonly labelled with fluorescent protein tags, which is not always feasible and was reported to cause artifacts. Organic DNA-binding dyes are free of these drawbacks, but they lack specificity to mitochondrial DNA. Here, considering physico-chemical properties of such dyes, we achieved preferential live-cell labelling of mitochondrial nucleoids by a nucleic acid staining dye SYBR Gold. It enabled time-lapse imaging of mitochondrial nucleoids by structured illumination microscopy and quantification of their motions.

 

PLoS ONE 13(9): e0203956.

DOI : 10.1371/journal.pone.0203956

Open Access : https://journals.plos.org/plosone/article/file?id=10.1371/journal.pone.0203956&type=printable

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