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High throughput microscopy

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https://www.readbyqxmd.com/read/28098273/high-definition-infrared-chemical-imaging-of-colorectal-tissue-using-a-spero-qcl-microscope
#1
B Bird, J Rowlette
Mid-infrared microscopy has become a key technique in the field of biomedical science and spectroscopy. This label-free, non-destructive technique permits the visualisation of a wide range of intrinsic biochemical markers in tissues, cells and biofluids by detection of the vibrational modes of the constituent molecules. Together, infrared microscopy and chemometrics is a widely accepted method that can distinguish healthy and diseased states with high accuracy. However, despite the exponential growth of the field and its research world-wide, several barriers currently exist for its full translation into the clinical sphere, namely sample throughput and data management...
January 18, 2017: Analyst
https://www.readbyqxmd.com/read/28096363/an-rna-based-signature-enables-high-specificity-detection-of-circulating-tumor-cells-in-hepatocellular-carcinoma
#2
Mark Kalinich, Irun Bhan, Tanya T Kwan, David T Miyamoto, Sarah Javaid, Joseph A LiCausi, John D Milner, Xin Hong, Lipika Goyal, Srinjoy Sil, Melissa Choz, Uyen Ho, Ravi Kapur, Alona Muzikansky, Huidan Zhang, David A Weitz, Lecia V Sequist, David P Ryan, Raymond T Chung, Andrew X Zhu, Kurt J Isselbacher, David T Ting, Mehmet Toner, Shyamala Maheswaran, Daniel A Haber
Circulating tumor cells (CTCs) are shed into the bloodstream by invasive cancers, but the difficulty inherent in identifying these rare cells by microscopy has precluded their routine use in monitoring or screening for cancer. We recently described a high-throughput microfluidic CTC-iChip, which efficiently depletes hematopoietic cells from blood specimens and enriches for CTCs with well-preserved RNA. Application of RNA-based digital PCR to detect CTC-derived signatures may thus enable highly accurate tissue lineage-based cancer detection in blood specimens...
January 17, 2017: Proceedings of the National Academy of Sciences of the United States of America
https://www.readbyqxmd.com/read/28095175/evaluation-of-the-droplet-microarray-platform-for-high-throughput-screening-of-suspension-cells
#3
Anna A Popova, Claire Depew, Katya Manuella Permana, Alexander Trubitsyn, Ravindra Peravali, Jorge Ángel González Ordiano, Markus Reischl, Pavel A Levkin
Phenotypic cell-based high-throughput screenings play a central role in drug discovery and toxicology. The main tendency in cell screenings is the increase of the throughput and decrease of reaction volume in order to accelerate the experiments, reduce the costs, and enable screenings of rare cells. Conventionally, cell-based assays are performed in microtiter plates, which exist in 96- to 1536-wells formats and cannot be further miniaturized. In addition, performing screenings of suspension cells is associated with risk of losing cell content during the staining procedures and incompatibility with high-content microscopy...
November 1, 2016: Journal of Laboratory Automation
https://www.readbyqxmd.com/read/28079286/design-synthesis-biological-evaluation-and-3d-qsar-analysis-of-podophyllotoxin-dioxazole-combination-as-tubulin-targeting-anticancer-agents
#4
Zi-Zhen Wang, Wen-Xue Sun, Xue Wang, Ya-Han Zhang, Han-Yue Qiu, Jin-Liang Qi, Yan-Jun Pang, Gui-Hua Lu, Xiao-Ming Wang, Fu-Gen Yu, Yong-Hua Yang
The advancement of cancer fighting drugs has never been a simple linear process. Those drugs design professionals begin to find inspiration from the nature after failing to find the ideal products by creative drugs design and high-throughput screening. To obtain new molecules for inhibiting tubulin, podophyllotoxin was adopted as the leading compound and 1,3,4-oxadiazole was brought in to the C-4 site of podophyllotoxin in this research. A series of seventeen podophyllotoxin derived esters have been achieved and then evaluated their antitumor activities against four different cancer cell lines: A549, MCF-7, HepG2 and HeLa...
January 12, 2017: Chemical Biology & Drug Design
https://www.readbyqxmd.com/read/28075531/three-dimensional-imaging-flow-cytometry-through-light-sheet-fluorescence-microscopy
#5
REVIEW
Emilio J Gualda, Hugo Pereira, Gabriel G Martins, Rui Gardner, Nuno Moreno
Flow cytometry is the tool of choice for high-speed acquisition and analysis of large cell populations, with the tradeoff of lacking intracellular spatial information. Although in the last decades flow cytometry systems that can actually acquire two-dimensional spatial information were developed, some of the limitations remained though, namely constrains related to sample size and lack of depth or dynamic information. The combination of fluidics and light-sheet illumination has the potential to address these limitations...
January 11, 2017: Cytometry. Part A: the Journal of the International Society for Analytical Cytology
https://www.readbyqxmd.com/read/28075486/highly-expandable-human-ips-cell-derived-neural-progenitor-cells-npc-and-neurons-for-central-nervous-system-disease-modeling-and-high-throughput-screening
#6
Chialin Cheng, Daniel M Fass, Kat Folz-Donahue, Marcy E MacDonald, Stephen J Haggarty
Reprogramming of human somatic cells into induced pluripotent stem (iPS) cells has greatly expanded the set of research tools available to investigate the molecular and cellular mechanisms underlying central nervous system (CNS) disorders. Realizing the promise of iPS cell technology for the identification of novel therapeutic targets and for high-throughput drug screening requires implementation of methods for the large-scale production of defined CNS cell types. Here we describe a protocol for generating stable, highly expandable, iPS cell-derived CNS neural progenitor cells (NPC) using multi-dimensional fluorescence activated cell sorting (FACS) to purify NPC defined by cell surface markers...
January 11, 2017: Current Protocols in Human Genetics
https://www.readbyqxmd.com/read/28073262/fluorescence-self-quenching-from-reporter-dyes-informs-on-the-structural-properties-of-amyloid-clusters-formed-in-vitro-and-in-cells
#7
WeiYue Chen, Laurence J Young, Meng Lu, Alessio Zaccone, Florian Ströhl, Na Yu, Gabriele S Kaminski Schierle, Clemens F Kaminski
The characterization of the aggregation kinetics of protein amyloids and the structural properties of the ensuing aggregates are vital in the study of the pathogenesis of many neurodegenerative diseases and the discovery of therapeutic targets. In this article, we show that the fluorescence lifetime of synthetic dyes covalently attached to amyloid proteins informs on the structural properties of amyloid clusters formed both in vitro and in cells. We demonstrate that the mechanism behind such a "lifetime sensor" of protein aggregation is based on fluorescence self-quenching and that it offers a good dynamic range to report on various stages of aggregation without significantly perturbing the process under investigation...
January 11, 2017: Nano Letters
https://www.readbyqxmd.com/read/28060301/cryosectioning-of-contiguous-regions-of-a-single-mouse-skeletal-muscle-for-gene-expression-and-histological-analyses
#8
Aaron M Beedle
With this method, consecutive cryosections are collected to enable both microscopy applications for tissue histology and enrichment of RNA for gene expression using adjacent regions from a single mouse skeletal muscle. Typically, it is challenging to achieve adequate homogenization of small skeletal muscle samples because buffer volumes may be too low for efficient grinding applications, yet without sufficient mechanical disruption, the dense tissue architecture of muscle limits penetration of buffer reagents, ultimately causing low RNA yield...
December 12, 2016: Journal of Visualized Experiments: JoVE
https://www.readbyqxmd.com/read/28060274/gap-junctional-intercellular-communication-a-functional-biomarker-to-assess-adverse-effects-of-toxicants-and-toxins-and-health-benefits-of-natural-products
#9
Brad L Upham, Iva Sovadinová, Pavel Babica
This protocol describes a scalpel loading-fluorescent dye transfer (SL-DT) technique that measures intercellular communication through gap junction channels, which is a major intercellular process by which tissue homeostasis is maintained. Interruption of gap junctional intercellular communication (GJIC) by toxicants, toxins, drugs, etc. has been linked to numerous adverse health effects. Many genetic-based human diseases have been linked to mutations in gap junction genes. The SL-DT technique is a simple functional assay for the simultaneous assessment of GJIC in a large population of cells...
December 25, 2016: Journal of Visualized Experiments: JoVE
https://www.readbyqxmd.com/read/28059233/cytopathological-image-analysis-using-deep-learning-networks-in-microfluidic-microscopy
#10
G Gopakumar, K Hari Babu, Deepak Mishra, Sai Siva Gorthi, Gorthi R K Sai Subrahmanyam
Cytopathologic testing is one of the most critical steps in the diagnosis of diseases, including cancer. However, the task is laborious and demands skill. Associated high cost and low throughput drew considerable interest in automating the testing process. Several neural network architectures were designed to provide human expertise to machines. In this paper, we explore and propose the feasibility of using deep-learning networks for cytopathologic analysis by performing the classification of three important unlabeled, unstained leukemia cell lines (K562, MOLT, and HL60)...
January 1, 2017: Journal of the Optical Society of America. A, Optics, Image Science, and Vision
https://www.readbyqxmd.com/read/28059116/further-characterization-of-maize-chlorotic-mottle-virus-and-its-synergistic-interaction-with-sugarcane-mosaic-virus-in-maize
#11
Qiang Wang, Chao Zhang, Chunyan Wang, Yajuan Qian, Zhenghe Li, Jian Hong, Xueping Zhou
Maize chlorotic mottle virus (MCMV) was first reported in maize in China in 2009. In this study we further analyzed the epidemiology of MCMV and corn lethal necrosis disease (CLND) in China. We determined that CLND observed in China was caused by co-infection of MCMV and sugarcane mosaic virus (SCMV). Phylogenetic analysis using four full-length MCMV cDNA sequences obtained in this study and the available MCMV sequences retrieved from GenBank indicated that Chinese MCMV isolates were derived from the same source...
January 6, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28057503/evaluation-of-cancer-cell-deformability-by-microcavity-array
#12
Tomoko Yoshino, Tsuyoshi Tanaka, Seita Nakamura, Ryo Negishi, Nozomi Shionoiri, Masahito Hosokawa, Tadashi Matsunaga
A cell entrapment device consisting of a microcavity array was used to analyze the deformability of MCF-10 human breast epithelial and MCF-7 human breast cancer cell lines by confocal laser scanning microscopy. Entrapment of up to 8 × 10(3) cells was achieved within 3 min. Protrusions were formed at the bottom surface of the array with a pore size of 3 μm. Protrusion length increased at higher filtration pressures and could be used to distinguish between MCF-7 and MCF-10 cells. These results indicate that our system is useful for high-throughput deformability analysis of cancer cells, which can provide insight into the mechanisms underlying tumor cell malignancy...
January 3, 2017: Analytical Biochemistry
https://www.readbyqxmd.com/read/28029779/hydrogel-droplet-microfluidics-for-high-throughput-single-molecule-cell-analysis
#13
Zhi Zhu, Chaoyong James Yang
Heterogeneity among individual molecules and cells has posed significant challenges to traditional bulk assays, due to the assumption of average behavior, which would lose important biological information in heterogeneity and result in a misleading interpretation. Single molecule/cell analysis has become an important and emerging field in biological and biomedical research for insights into heterogeneity between large populations at high resolution. Compared with the ensemble bulk method, single molecule/cell analysis explores the information on time trajectories, conformational states, and interactions of individual molecules/cells, all key factors in the study of chemical and biological reaction pathways...
December 28, 2016: Accounts of Chemical Research
https://www.readbyqxmd.com/read/28009439/sudan-black-a-fast-easy-and-non-toxic-method-to-assess-myelin-repair-in-demyelinating-diseases
#14
Benjamin V Ineichen, Oliver Weinmann, Nicolas Good, Patricia Plattner, Carla Wicki, Elisabeth J Rushing, Michael Linnebank, Martin E Schwab
AIMS: The search for novel drugs that enhance myelin repair in entities such as multiple sclerosis has top priority in neurological research, not least because remyelination can hinder further neurodegeneration in neuro-inflammatory conditions. Recently, several new compounds with the potential to boost remyelination have been identified using high-throughput in vitro screening methods. However, assessing their potential to enhance remyelination in vivo using plastic embedded semi-thin sections or electron microscopy, even though being the gold standard for assessing remyelination, is toxic, extremely time-consuming, and expensive...
December 23, 2016: Neuropathology and Applied Neurobiology
https://www.readbyqxmd.com/read/27995336/characterization-and-complete-genome-sequence-of-a-panicovirus-from-bermuda-grass-by-high-throughput-sequencing
#15
Muhammad N Tahir, Ben Lockhart, Samuel Grinstead, Dimitre Mollov
Bermuda grass samples were examined by transmission electron microscopy and 28-30 nm spherical virus particles were observed. Total RNA from these plants was subjected to high-throughput sequencing (HTS). The nearly full genome sequence of a panicovirus was identified from one HTS scaffold. Sanger sequencing was used to confirm the HTS results and complete the genome sequence of 4404 nt. This virus was provisionally named Bermuda grass latent virus (BGLV). Its predicted open reading frames follow the typical arrangement of the genus Panicovirus...
December 19, 2016: Archives of Virology
https://www.readbyqxmd.com/read/27982581/comprehensive-landscape-of-nrf2-and-p53-pathway-activation-dynamics-by-oxidative-stress-and-dna-damage
#16
Steven Hiemstra, Marije Niemeijer, Esmee Koedoot, Steven Wink, Janna E Klip, Matthijs Vlasveld, Elisabeth de Zeeuw, Bram van Os, Andrew White, Bob van de Water
A quantitative dynamics pathway map of the Nrf2-mediated oxidative stress response and p53-related DNA damage response pathways as well as the cross-talk between these pathways has not systematically been defined. To allow the dynamic single cell evaluation of these pathways, we have used BAC-GFP recombineering to tag for each pathway's three key components: for the oxidative stress response, Keap1-GFP, Nrf2-GFP, and Srxn1-GFP; for the DNA damage response, 53bp1-GFP, p53-GFP, and p21-GFP. The dynamic activation of these individual components was assessed using quantitative high throughput confocal microscopy after treatment with a broad concentration range of diethyl maleate (DEM; to induce oxidative stress) and etoposide (to induce DNA damage)...
December 16, 2016: Chemical Research in Toxicology
https://www.readbyqxmd.com/read/27981718/a-redox-activatable-fluorescent-sensor-for-the-high-throughput-quantification-of-cytosolic-delivery-of-macromolecules
#17
Zhaohui Wang, Min Luo, Chengqiong Mao, Qi Wei, Tian Zhao, Yang Li, Gang Huang, Jinming Gao
Efficient delivery of biomacromolecules (e.g., proteins, nucleic acids) into cell cytosol remains a critical challenge for the development of macromolecular therapeutics or diagnostics. To date, most common approaches to assess cytosolic delivery rely on fluorescent labeling of macromolecules with an "always on" reporter and subcellular imaging of endolysosomal escape by confocal microscopy. This strategy is limited by poor signal-to-noise ratio and only offers low throughput, qualitative information. Herein we describe a quantitative redox-activatable sensor (qRAS) for the real-time monitoring of cytosolic delivery of macromolecules...
December 16, 2016: Angewandte Chemie
https://www.readbyqxmd.com/read/27980506/atomistic-modelling-of-scattering-data-in-the-collaborative-computational-project-for-small-angle-scattering-ccp-sas
#18
Stephen J Perkins, David W Wright, Hailiang Zhang, Emre H Brookes, Jianhan Chen, Thomas C Irving, Susan Krueger, David J Barlow, Karen J Edler, David J Scott, Nicholas J Terrill, Stephen M King, Paul D Butler, Joseph E Curtis
The capabilities of current computer simulations provide a unique opportunity to model small-angle scattering (SAS) data at the atomistic level, and to include other structural constraints ranging from molecular and atomistic energetics to crystallography, electron microscopy and NMR. This extends the capabilities of solution scattering and provides deeper insights into the physics and chemistry of the systems studied. Realizing this potential, however, requires integrating the experimental data with a new generation of modelling software...
December 1, 2016: Journal of Applied Crystallography
https://www.readbyqxmd.com/read/27978709/a-malaria-transmission-blocking-usnic-acid-derivative-prevents-plasmodium-zygote-to-ookinete-maturation-in-the-mosquito-midgut
#19
Rebecca Pastrana-Mena, Derrick K Mathias, Michael Delves, Krithika Rajaram, Jonas G King, Rebecca Yee, Beatrice Trucchi, Luisella Verotta, Rhoel R Dinglasan
The evolution of drug resistance is a recurrent problem that has plagued efforts to treat and control malaria. Recent emergence of artemisinin resistance in Southeast Asia underscores the need to develop novel antimalarials and identify new targetable pathways in Plasmodium parasites. Transmission-blocking approaches, which typically target gametocytes in the host bloodstream or parasite stages in the mosquito gut, are recognized collectively as a strategy that when used in combination with antimalarials that target erythrocytic stages will not only cure malaria but will also prevent subsequent transmission...
December 16, 2016: ACS Chemical Biology
https://www.readbyqxmd.com/read/27975246/detection-and-quantification-methods-for-fibrillar-products-of-in-vitro-tau-aggregation-assays
#20
Niki Nanavaty, Lauren Lin, Samantha H Hinckley, Jeff Kuret
Alzheimer's disease is characterized in part by the intracellular misfolding and aggregation of tau protein. The aggregates, which range in size from small oligomers to large filaments, are markers for disease diagnosis and staging, potential vectors for disease propagation, and candidate sources of neurotoxicity. Here we present protocols for synthesizing large tau aggregates characterized by filamentous morphology and cross-β-sheet structure from monomeric full-length tau precursors in vitro. We also describe their detection and quantification through thioflavin dye binding, filter trap, and transmission electron microscopy methods...
2017: Methods in Molecular Biology
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