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High throughput microscopy

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https://www.readbyqxmd.com/read/27911388/high-throughput-real-time-dual-readout-testing-of-intracellular-antimicrobial-activity-and-eukaryotic-cell-cytotoxicity
#1
Lucius Chiaraviglio, Yoon-Suk Kang, James E Kirby
Traditional measures of intracellular antimicrobial activity and eukaryotic cell cytotoxicity rely on endpoint assays. Such endpoint assays require several additional experimental steps prior to readout, such as cell lysis, colony forming unit determination, or reagent addition. When performing thousands of assays, for example, during high-throughput screening, the downstream effort required for these types of assays is considerable. Therefore, to facilitate high-throughput antimicrobial discovery, we developed a real-time assay to simultaneously identify inhibitors of intracellular bacterial growth and assess eukaryotic cell cytotoxicity...
November 16, 2016: Journal of Visualized Experiments: JoVE
https://www.readbyqxmd.com/read/27909234/a-high-throughput-screening-strategy-for-development-of-rnf8-ubc13-protein-protein-interaction-inhibitors
#2
Elisabeth Weber, Ina Rothenaigner, Stefanie Brandner, Kamyar Hadian, Kenji Schorpp
The ubiquitin-proteasome system plays an essential role in a broad range of cellular signaling pathways. Ubiquitination is a posttranslational protein modification that involves the action of an enzymatic cascade (E1, E2, and E3 enzymes) for the covalent attachment of ubiquitin to target proteins. The emerging knowledge of the molecular mechanisms and correlation of deregulation of the ubiquitin system in human diseases is uncovering new opportunities for therapeutics development. The E3 ligase RNF8 acts in cooperation with the heterodimeric E2 enzyme Ubc13/Uev1a to generate ubiquitin conjugates at the sides of DNA double-strand breaks, and recent findings suggest RNF8 as a potential therapeutic target for the treatment of breast cancer...
December 1, 2016: Journal of Biomolecular Screening
https://www.readbyqxmd.com/read/27905503/advancing-ovarian-folliculometry-with-selective-plane-illumination-microscopy
#3
Hsiao-Chun Amy Lin, Rahul Dutta, Subhamoy Mandal, Alexander Kind, Angelika Schnieke, Daniel Razansky
Determination of ovarian status and follicle monitoring are common methods of diagnosing female infertility. We evaluated the suitability of selective plane illumination microscopy (SPIM) for the study of ovarian follicles. The large field of view and fast acquisition speed of our SPIM system enables rendering of volumetric image stacks from intact whole porcine ovarian follicles, clearly visualizing follicular features including follicle volume and average diameter (70 μm-2.5 mm), their spherical asymmetry parameters, size of developing cumulus oophorus complexes (40 μm-110 μm), and follicular wall thickness (90 μm-120 μm)...
December 1, 2016: Scientific Reports
https://www.readbyqxmd.com/read/27903742/p2y6-receptor-antagonist-mrs2578-inhibits-neutrophil-activation-and-aggregated-neutrophil-extracellular-trap-formation-induced-by-gout-associated-monosodium-urate-crystals
#4
Payel Sil, Craig P Hayes, Barbara J Reaves, Patrick Breen, Shannon Quinn, Jeremy Sokolove, Balázs Rada
Human neutrophils (polymorphonuclear leukocytes [PMNs]) generate inflammatory responses within the joints of gout patients upon encountering monosodium urate (MSU) crystals. Neutrophil extracellular traps (NETs) are found abundantly in the synovial fluid of gout patients. The detailed mechanism of MSU crystal-induced NET formation remains unknown. Our goal was to shed light on possible roles of purinergic signaling and neutrophil migration in mediating NET formation induced by MSU crystals. Interaction of human neutrophils with MSU crystals was evaluated by high-throughput live imaging using confocal microscopy...
November 30, 2016: Journal of Immunology: Official Journal of the American Association of Immunologists
https://www.readbyqxmd.com/read/27901048/sparsity-based-multi-height-phase-recovery-in-holographic-microscopy
#5
Yair Rivenson, Yichen Wu, Hongda Wang, Yibo Zhang, Alborz Feizi, Aydogan Ozcan
High-resolution imaging of densely connected samples such as pathology slides using digital in-line holographic microscopy requires the acquisition of several holograms, e.g., at >6-8 different sample-to-sensor distances, to achieve robust phase recovery and coherent imaging of specimen. Reducing the number of these holographic measurements would normally result in reconstruction artifacts and loss of image quality, which would be detrimental especially for biomedical and diagnostics-related applications...
November 30, 2016: Scientific Reports
https://www.readbyqxmd.com/read/27896026/jicbioimage-a-tool-for-automated-and-reproducible-bioimage-analysis
#6
Tjelvar S G Olsson, Matthew Hartley
There has been steady improvement in methods for capturing bioimages. However analysing these images still remains a challenge. The Python programming language provides a powerful and flexible environment for scientific computation. It has a wide range of supporting libraries for image processing but lacks native support for common bioimage formats, and requires specific code to be written to ensure that suitable audit trails are generated and analyses are reproducible. Here we describe the development of a Python tool that: (1) allows users to quickly view and explore microscopy data; (2) generate reproducible analyses, encoding a complete history of image transformations from raw data to final result; and (3) scale up analyses from initial exploration to high throughput processing pipelines, with a minimal amount of extra effort...
2016: PeerJ
https://www.readbyqxmd.com/read/27896006/ultrahigh-speed-phase-sensitive-full-field-interferometric-confocal-microscopy-for-quantitative-microscale-physiology
#7
Ikbal Sencan, Brendan K Huang, Yong Bian, Emily Mis, Mustafa K Khokha, Hui Cao, Michael Choma
We developed ultra-high-speed, phase-sensitive, full-field reflection interferometric confocal microscopy (FFICM) for the quantitative characterization of in vivo microscale biological motions and flows. We demonstrated 2D frame rates in excess of 1 kHz and pixel throughput rates up to 125 MHz. These fast FFICM frame rates were enabled by the use of a low spatial coherence, high-power laser source. Specifically, we used a dense vertical cavity surface emitting laser (VCSEL) array that synthesized low spatial coherence light through a large number of narrowband, mutually-incoherent emitters...
November 1, 2016: Biomedical Optics Express
https://www.readbyqxmd.com/read/27881953/connectomic-analysis-of-brain-networks-novel-techniques-and-future-directions
#8
REVIEW
J Leonie Cazemier, Francisco Clascá, Paul H E Tiesinga
Brain networks, localized or brain-wide, exist only at the cellular level, i.e., between specific pre- and post-synaptic neurons, which are connected through functionally diverse synapses located at specific points of their cell membranes. "Connectomics" is the emerging subfield of neuroanatomy explicitly aimed at elucidating the wiring of brain networks with cellular resolution and a quantified accuracy. Such data are indispensable for realistic modeling of brain circuitry and function. A connectomic analysis, therefore, needs to identify and measure the soma, dendrites, axonal path, and branching patterns together with the synapses and gap junctions of the neurons involved in any given brain circuit or network...
2016: Frontiers in Neuroanatomy
https://www.readbyqxmd.com/read/27876275/development-of-leishmania-donovani-stably-expressing-dsred-for-flow-cytometry-based-drug-screening-using-chalcone-thiazolyl-hydrazone-as-a-new-antileishmanial-target
#9
Anil Kumar Jaiswal, K Bhaskara Rao, Pragati Kushwaha, Keerti Rawat, Ram K Modukuri, Prashant Khare, Sumit Joshi, Shikha Mishra, Ambak Rai, Koneni V Sashidhara, Anuradha Dube
Green fluorescent protein produces significant fluorescence and is extremely stable, however its excitation maximum is close to the ultraviolet range and thus can damage living cells. Hence, Leishmania donovani stably expressing DsRed were developed and their suitability for flow cytometry-based antileishmanial screening was assessed by evaluating the efficacies of standard drugs as well as newly synthesised chalcone thiazolyl-hydrazone compounds. The DsRed gene was successfully integrated at the 18S rRNA locus of L...
November 2, 2016: International Journal of Antimicrobial Agents
https://www.readbyqxmd.com/read/27873203/optical-microscopy-for-detecting-binding-on-small-molecule-microarrays
#10
Si Ying Png, Sherlyn Xiang Bing Seah, Zhiqiang Gao
Small molecule microarrays (SMMs) have revolutionized parallel screening of multiple biological interactions. SMMs are a high-throughput technique for simultaneous detection of up to thousands of biological interactions. Conventional detection technique employed in SMMs is fluorescence spectroscopy. However, the possibility of fluorescent labels affecting the binding properties of small molecules drives the development of label-free screening techniques. Here, we describe two label-free detection techniques using optical microscopy for the detection of binding on SMMs...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/27873202/label-free-sensing-on-microarrays
#11
Yung-Shin Sun
Microarrays of biological molecules such as DNAs, proteins, carbohydrates, and small molecules provide a high-throughput platform for screening tens of thousands of biomolecular interactions simultaneously, facilitating the functional characterization of these biomolecules in areas of genomics, proteomics, glycomics, and cytomics. Routinely, analysis of binding reactions between solution-phased probes and surface-immobilized targets involves some kinds of fluorescence-based detection methods. Even though these methods have advantages of high sensitivity and wide dynamic range, labeling probes and/or targets inevitably changes their innate properties and in turn affects probe-target interactions in often uncharacterized ways...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/27870847/creating-high-resolution-multiscale-maps-of-human-tissue-using-multi-beam-sem
#12
André F Pereira, Daniel J Hageman, Tomasz Garbowski, Christof Riedesel, Ulf Knothe, Dirk Zeidler, Melissa L Knothe Tate
Multi-beam scanning electron microscopy (mSEM) enables high-throughput, nano-resolution imaging of macroscopic tissue samples, providing an unprecedented means for structure-function characterization of biological tissues and their cellular inhabitants, seamlessly across multiple length scales. Here we describe computational methods to reconstruct and navigate a multitude of high-resolution mSEM images of the human hip. We calculated cross-correlation shift vectors between overlapping images and used a mass-spring-damper model for optimal global registration...
November 2016: PLoS Computational Biology
https://www.readbyqxmd.com/read/27869673/3d-visualization-of-developmental-toxicity-of-2-4-6-trinitrotoluene-in-zebrafish-embryogenesis-using-light-sheet-microscopy
#13
Juneyong Eum, Jina Kwak, Hee Joung Kim, Seoyoung Ki, Kooyeon Lee, Ahmed A Raslan, Ok Kyu Park, Md Ashraf Uddin Chowdhury, Song Her, Yun Kee, Seung-Hae Kwon, Byung Joon Hwang
Environmental contamination by trinitrotoluene is of global concern due to its widespread use in military ordnance and commercial explosives. Despite known long-term persistence in groundwater and soil, the toxicological profile of trinitrotoluene and other explosive wastes have not been systematically measured using in vivo biological assays. Zebrafish embryos are ideal model vertebrates for high-throughput toxicity screening and live in vivo imaging due to their small size and transparency during embryogenesis...
November 17, 2016: International Journal of Molecular Sciences
https://www.readbyqxmd.com/read/27864781/isolation-identification-and-high-throughput-screening-of-neutral-lipid-producing-indigenous-microalgae-from-south-african-aquatic-habitats
#14
S T Gumbi, B M Majeke, A O Olaniran, T Mutanda
Exploring indigenous microalgae capable of producing significant amounts of neutral lipids through high-throughput screening is crucial for sustainable biodiesel production. In this study, 31 indigenous microalgal strains were isolated from diverse aquatic habitats in KwaZulu-Natal, South Africa. Eight superior lipid-producing strains were selected for further analysis, based on Nile red fluorescence microscopy screening. The microalgal isolates were identified to belong to the genera Chlorella, Neochloris and Chlamydomonas via morpho-taxonomic and molecular approach by 18S rRNA gene sequencing...
November 18, 2016: Applied Biochemistry and Biotechnology
https://www.readbyqxmd.com/read/27858020/a-protocol-for-rapid-label-free-histochemical-imaging-of-fibrotic-liver
#15
B Bird, J Rowlette
Mid-infrared microscopy is a non-destructive, quantitative and label-free spectroscopic imaging technique that, as a result of recent instrument advancements, is now at the point of enabling high-throughput automated biochemical screening of whole histology samples. Currently the mid-infrared field is undergoing a paradigm shift that has not been seen since the introduction of scanning Fourier Transform interferometric spectrometers. The latest mid-infrared microscopes are powered by tunable quantum cascade laser (QCL) sources which offer a number of advantages including measurement protocol flexibility, ease-of-use and a greatly enhanced data acquisition speed at high spectral and spatial resolution...
November 18, 2016: Analyst
https://www.readbyqxmd.com/read/27846239/high-throughput-accurate-single-cell-screening-of-euglena-gracilis-with-fluorescence-assisted-optofluidic-time-stretch-microscopy
#16
Baoshan Guo, Cheng Lei, Takuro Ito, Yiyue Jiang, Yasuyuki Ozeki, Keisuke Goda
The development of reliable, sustainable, and economical sources of alternative fuels is an important, but challenging goal for the world. As an alternative to liquid fossil fuels, algal biofuel is expected to play a key role in alleviating global warming since algae absorb atmospheric CO2 via photosynthesis. Among various algae for fuel production, Euglena gracilis is an attractive microalgal species as it is known to produce wax ester (good for biodiesel and aviation fuel) within lipid droplets. To date, while there exist many techniques for inducing microalgal cells to produce and accumulate lipid with high efficiency, few analytical methods are available for characterizing a population of such lipid-accumulated microalgae including E...
2016: PloS One
https://www.readbyqxmd.com/read/27830849/review-imaging-technologies-for-flow-cytometry
#17
REVIEW
Yuanyuan Han, Yi Gu, Alex Ce Zhang, Yu-Hwa Lo
High-throughput single cell imaging is a critical enabling and driving technology in molecular and cellular biology, biotechnology, medicine and related areas. Imaging flow cytometry combines the single-cell imaging capabilities of microscopy with the high-throughput capabilities of conventional flow cytometry. Recent advances in imaging flow cytometry are remarkably revolutionizing single-cell analysis. This article describes recent imaging flow cytometry technologies and their challenges.
November 10, 2016: Lab on a Chip
https://www.readbyqxmd.com/read/27827454/novel-high-throughput-approach-for-purification-of-infectious-virions
#18
Kevin T James, Brad Cooney, Kate Agopsowicz, Mary Ann Trevors, Adil Mohamed, Don Stoltz, Mary Hitt, Maya Shmulevitz
Viruses are extensively studied as pathogens and exploited as molecular tools and therapeutic agents. Existing methods to purify viruses such as gradient ultracentrifugation or chromatography have limitations, for example demand for technical expertise or specialized equipment, high time consumption, and restricted capacity. Our laboratory explores mutations in oncolytic reovirus that could improve oncolytic activity, and makes routine use of numerous virus variants, genome reassortants, and reverse engineered mutants...
November 9, 2016: Scientific Reports
https://www.readbyqxmd.com/read/27823959/sfida-automation-yields-sub-femtomolar-limit-of-detection-for-a%C3%AE-aggregates-in-body-fluids
#19
Yvonne Herrmann, Andreas Kulawik, Katja Kühbach, Maren Hülsemann, Luriano Peters, Tuyen Bujnicki, Kateryna Kravchenko, Christina Linnartz, Johannes Willbold, Christian Zafiu, Oliver Bannach, Dieter Willbold
OBJECTIVES: Alzheimer's disease (AD) is a neurodegenerative disorder with yet non-existent therapeutic and limited diagnostic options. Reliable biomarker-based AD diagnostics are of utmost importance for the development and application of therapeutic substances. We have previously introduced a platform technology designated 'sFIDA' for the quantitation of amyloid β peptide (Aβ) aggregates as AD biomarker. In this study we implemented the sFIDA assay on an automated platform to enhance robustness and performance of the assay...
November 5, 2016: Clinical Biochemistry
https://www.readbyqxmd.com/read/27822856/in-situ-single-molecule-rna-genotyping-using-padlock-probes-and-rolling-circle-amplification
#20
Tomasz Krzywkowski, Thomas Hauling, Mats Nilsson
Present-day techniques allow for massively parallel and high-throughput characterization of the somatic mutation status of samples. Most of these assays rely on whole specimen extracts, where heterogeneous spatial context of the specimen is lost. This chapter describes an up-to-date protocol for multiplexed, in situ genotyping of RNA in preserved tissue and cell lines, using padlock probes and rolling circle amplification. The presented approach allows for automated quantification of mRNA expression and mutation status, in single cells or in designated specimen areas...
2017: Methods in Molecular Biology
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