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Journal of Biomolecular Screening

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https://www.readbyqxmd.com/read/27932699/a-systematic-investigation-of-the-best-buffers-for-use-in-screening-by-maldi-mass-spectrometry
#1
Jessica Chandler, Carl Haslam, Neil Hardy, Melanie Leveridge, Peter Marshall
Matrix-assisted laser desorption/ionization-mass spectrometry (MALDI-MS) offers a label-free alternative for the screening of biochemical targets in both 1536- and 6144-assay formats, as well as potentially providing increased sensitivity, reproducibility, and the simultaneous detection of multiple assay components within a specified m/z range. Ion suppression effects are one of the principal limitations reported for MS analysis. Within MALDI-MS screening, it has been identified that certain biochemical components incorporated into the assay (e...
December 8, 2016: Journal of Biomolecular Screening
https://www.readbyqxmd.com/read/27932697/htra3-isoform-specific-elisas-for-early-detection-of-preeclampsia
#2
Yao Wang, Ying Li, Jonathan Hyett, Fabricio da Silva Costa, Guiying Nie
Preeclampsia is a serious disorder of human pregnancy occurring after 20 weeks of gestation. It can be divided into subtypes of early onset (<34 weeks of gestation) and late onset (>34 weeks). Presymptomatic detection to identify those at high risk is important for managing this disease. HtrA3, a serine protease with high expression in the developing placenta, exists in long (HtrA3-L) and short (HtrA3-S) isoforms. They are identical, except HtrA3-S lacks the C-terminal PDZ domain. We have previously shown by Western blot analysis that serum HtrA3 levels at the end of the first trimester are significantly higher in women who later develop preeclampsia than in controls...
December 8, 2016: Journal of Biomolecular Screening
https://www.readbyqxmd.com/read/28139961/functional-comparison-of-neuronal-cells-differentiated-from-human-induced-pluripotent-stem-cell-derived-neural-stem-cells-under-different-oxygen-and-medium-conditions
#3
Kazuto Yamazaki, Kazuyuki Fukushima, Michiko Sugawara, Yoshikuni Tabata, Yoichi Imaizumi, Yasuharu Ishihara, Masashi Ito, Kappei Tsukahara, Jun Kohyama, Hideyuki Okano
Because neurons are difficult to obtain from humans, generating functional neurons from human induced pluripotent stem cells (hiPSCs) is important for establishing physiological or disease-relevant screening systems for drug discovery. To examine the culture conditions leading to efficient differentiation of functional neural cells, we investigated the effects of oxygen stress (2% or 20% O2) and differentiation medium (DMEM/F12:Neurobasal-based [DN] or commercial [PhoenixSongs Biologicals; PS]) on the expression of genes related to neural differentiation, glutamate receptor function, and the formation of networks of neurons differentiated from hiPSCs (201B7) via long-term self-renewing neuroepithelial-like stem (lt-NES) cells...
December 2016: Journal of Biomolecular Screening
https://www.readbyqxmd.com/read/28139960/development-of-an-in-vitro-model-to-screen-cyp1b1-targeted-anticancer-prodrugs
#4
Zhiying Wang, Yao Chen, Laura M Drbohlav, Judy Qiju Wu, Michael Zhuo Wang
Cytochrome P450 1B1 (CYP1B1) is an anticancer therapeutic target due to its overexpression in a number of steroid hormone-related cancers. One anticancer drug discovery strategy is to develop prodrugs specifically activated by CYP1B1 in malignant tissues to cytotoxic metabolites. Here, we aimed to develop an in vitro screening model for CYP1B1-targeted anticancer prodrugs using the KLE human endometrial carcinoma cell line. KLE cells demonstrated superior stability of CYP1B1 expression relative to transiently transfected cells and did not express any appreciable amount of cognate CYP1A1 or CYP1A2, which would have compromised the specificity of the screening assay...
December 2016: Journal of Biomolecular Screening
https://www.readbyqxmd.com/read/28027450/enabling-1536-well-high-throughput-cell-based-screening-through-the-application-of-novel-centrifugal-plate-washing
#5
Sinead Knight, Helen Plant, Lisa McWilliams, David Murray, Rebecca Dixon-Steele, Anet Varghese, Paul Harper, Anna Ramne, Paula McArdle, Susanna Engberg, Neil Bennett, Carolyn Blackett, Mark Wigglesworth
Cell-based assays have long been important within hit discovery paradigms; however, improving the disease relevance of the assay system can positively affect the translation of small-molecule drug discovery, especially if adopted in the initial hit identification assay. Consequently, there is an increasing need for disease-relevant assay systems capable of running at large scale, including the use of induced pluripotent stem cells and donor-derived primary cells. Major hurdles to adopting these assays for high-throughput screening are the cost, availability of cells, and complex protocols...
December 1, 2016: Journal of Biomolecular Screening
https://www.readbyqxmd.com/read/28027448/induced-pluripotent-hd-monkey-stem-cells-derived-neural-cells-for-drug-discovery
#6
Tanut Kunkanjanawan, Richard Carter, Kwan-Sung Ahn, Jinjing Yang, Rangsun Parnpai, Anthony W S Chan
Huntington's disease (HD) is a neurodegenerative disease caused by an expansion of CAG trinucleotide repeat (polyglutamine [polyQ]) in the huntingtin ( HTT) gene, which leads to the formation of mutant HTT (mHTT) protein aggregates. In the nervous system, an accumulation of mHTT protein results in glutamate-mediated excitotoxicity, proteosome instability, and apoptosis. Although HD pathogenesis has been extensively studied, effective treatment of HD has yet to be developed. Therapeutic discovery research in HD has been reported using yeast, cells derived from transgenic animal models and HD patients, and induced pluripotent stem cells from patients...
December 1, 2016: Journal of Biomolecular Screening
https://www.readbyqxmd.com/read/27647668/one-step-seeding-of-neural-stem-cells-with-vitronectin-supplemented-medium-for-high-throughput-screening-assays
#7
Sheng Dai, Rong Li, Yan Long, Steve Titus, Jinghua Zhao, Ruili Huang, Menghang Xia, Wei Zheng
Human neuronal cells differentiated from induced pluripotent cells have emerged as a new model system for the study of disease pathophysiology and evaluation of drug efficacy. Differentiated neuronal cells are more similar in genetics and biological content to human brain cells than other animal disease models. However, culture of neuronal cells in assay plates requires a labor-intensive procedure of plate precoating, hampering its applications in high-throughput screening (HTS). We developed a simplified method with one-step seeding of neural stem cells in assay plates by supplementing the medium with a recombinant human vitronectin (VTN), thus avoiding plate precoating...
December 2016: Journal of Biomolecular Screening
https://www.readbyqxmd.com/read/27269812/a-platform-to-enable-the-pharmacological-profiling-of-small-molecules-in-gel-based-electrophoretic-mobility-shift-assays
#8
Timothy L Foley, Dorjbal Dorjsuren, Thomas S Dexheimer, Michael D Burkart, William C Wight, Anton Simeonov
We describe a polyacrylamide gel casting cassette that overcomes limitations of commercially available gel electrophoresis equipment. This apparatus molds a single polyacrylamide gel that can evaluate more than 200 samples in parallel, is loaded with a multichannel pipettor, and is flexible with respect to composition of the separating matrix. We demonstrate its use to characterize inhibitors of enzymes that modify protein and nucleic acid substrates. Throughputs of greater than 1000 samples per day were achieved when this system was paired with a quantitative laser-based imaging system, yielding data of remarkable quality...
December 2016: Journal of Biomolecular Screening
https://www.readbyqxmd.com/read/27872201/identification-of-chemical-compounds-that-inhibit-protein-synthesis-in-pseudomonas-aeruginosa
#9
Stephanie O Palmer, Yanmei Hu, Megan Keniry, James M Bullard
Four inhibitory compounds were identified using a poly-uridylic acid (polyU) mRNA-directed aminoacylation/translation (A/T) protein synthesis system composed of phenylalanyl-tRNA synthetases (PheRS), ribosomes, and ribosomal factors from Pseudomonas aeruginosa in an in vitro screen of a synthetic compound library. The compounds were specific for inhibition of bacterial protein synthesis. In enzymatic assays, the compounds inhibited protein synthesis with IC50 values ranging from 20 to 60 μM. Minimum inhibitory concentrations (MICs) were determined in cultures for a panel of pathogenic organisms, including Enterococcus faecalis, Escherichia coli, Haemophilus influenzae, P...
November 21, 2016: Journal of Biomolecular Screening
https://www.readbyqxmd.com/read/27810952/high-content-screening-of-raw-actinomycete-extracts-for-the-identification-of-antituberculosis-activities
#10
Jinyeong Heo, Jiyoun Nam, Jichan Jang, David Shum, Constantin Radu, Jinhua Cheng, Hanki Lee, Joo-Won Suh, Vincent Delorme
The feasibility and relevance of screening a library of raw actinomycete extracts (ECUM library) for the identification of antituberculosis activities was assessed on 11,088 extracts using a multiple-screening approach. Each extract was first tested at two concentrations against noninfected macrophages as a control, then against Mycobacterium tuberculosis growing in broth medium as well as infecting murine macrophages. The screening results indicated a library of good quality with an apparent low proportion of cytotoxic extracts...
November 3, 2016: Journal of Biomolecular Screening
https://www.readbyqxmd.com/read/27789755/a-genome-wide-arrayed-cdna-screen-to-identify-functional-modulators-of-%C3%AE-7-nicotinic-acetylcholine-receptors
#11
Elizabeth B Rex, Nikhil Shukla, Shenyan Gu, David Bredt, Daniel DiSepio
Cellular signaling is in part regulated by the composition and subcellular localization of a series of protein interactions that collectively form a signaling complex. Using the α7 nicotinic acetylcholine receptor (α7nAChR) as a proof-of-concept target, we developed a platform to identify functional modulators (or auxiliary proteins) of α7nAChR signaling. The Broad cDNA library was transiently cotransfected with α7nAChR cDNA in HEK293T cells in a high-throughput fashion. Using this approach in combination with a functional assay, we identified positive modulators of α7nAChR activity...
October 27, 2016: Journal of Biomolecular Screening
https://www.readbyqxmd.com/read/27789753/an-escherichia-coli-based-assay-to-assess-the-function-of-recombinant-human-hemichannels
#12
Srinivasan Krishnan, Mariana C Fiori, Ty E Whisenant, D Marien Cortes, Guillermo A Altenberg, Luis G Cuello
Connexins form the gap junctional channels that mediate cell-to-cell communication, and also form hemichannels present at the plasma membrane. Hemichannels are permeable to small hydrophilic compounds, including molecules involved in autocrine and paracrine signaling. An abnormal hemichannel opening causes or contributes to cell damage in common human disorders (e.g., cardiac infarct, cerebrovascular accidents, deafness, skin diseases, and cataracts) and is therefore a potential pharmacological target. The discovery of useful hemichannels inhibitors has been hampered in part by the lack of suitable high-throughput functional assays...
October 27, 2016: Journal of Biomolecular Screening
https://www.readbyqxmd.com/read/27729504/efficient-management-of-high-throughput-screening-libraries-with-savanah
#13
Markus List, Marlene Pedersen Elnegaard, Steffen Schmidt, Helle Christiansen, Qihua Tan, Jan Mollenhauer, Jan Baumbach
High-throughput screening (HTS) has become an indispensable tool for the pharmaceutical industry and for biomedical research. A high degree of automation allows for experiments in the range of a few hundred up to several hundred thousand to be performed in close succession. The basis for such screens are molecular libraries, that is, microtiter plates with solubilized reagents such as siRNAs, shRNAs, miRNA inhibitors or mimics, and sgRNAs, or small compounds, that is, drugs. These reagents are typically condensed to provide enough material for covering several screens...
October 11, 2016: Journal of Biomolecular Screening
https://www.readbyqxmd.com/read/27729503/evaluation-of-parameters-impacting-drug-susceptibility-in-intracellular-trypanosoma-cruzi-assay-protocols
#14
Gyongseon Yang, Nakyung Lee, Jean-Robert Ioset, Joo Hwan No
In order to understand the key parameters influencing drug susceptibility, different Trypanosoma cruzi assay protocols were evaluated using a comparative assay design. The assays compared in this study were an image-based intracellular T. cruzi assay quantified through an image-mining algorithm and an intracellular assay utilizing a β-galactosidase-expressing T. cruzi strain. Thirty-one reference compounds known to exhibit activities against intracellular T. cruzi were used as benchmarks. Initial comparison using EC50 values from two assays showed a very poor correlation, with an R(2) value of 0...
October 11, 2016: Journal of Biomolecular Screening
https://www.readbyqxmd.com/read/27650791/high-throughput-platform-for-identifying-molecular-factors-involved-in-phenotypic-stabilization-of-primary-human-hepatocytes-in-vitro
#15
Jing Shan, David J Logan, David E Root, Anne E Carpenter, Sangeeta N Bhatia
Liver disease is a leading cause of morbidity worldwide and treatment options are limited, with organ transplantation being the only form of definitive management. Cell-based therapies have long held promise as alternatives to whole-organ transplantation but have been hindered by the rapid loss of liver-specific functions over a period of days in cultured hepatocytes. Hypothesis-driven studies have identified a handful of factors that modulate hepatocyte functions in vitro, but our understanding of the mechanisms involved remains incomplete...
October 2016: Journal of Biomolecular Screening
https://www.readbyqxmd.com/read/27650790/slas-europe-high-content-screening-conference-in-dresden-a-glimpse-of-the-future
#16
EDITORIAL
Maria Montoya, Thierry Dorval, Marc Bickle
No abstract text is available yet for this article.
October 2016: Journal of Biomolecular Screening
https://www.readbyqxmd.com/read/27461835/development-of-a-high-throughput-gene-expression-screen-for-modulators-of-ras-mapk-signaling-in-a-mutant-ras-cellular-context
#17
Bryan Severyn, Thi Nguyen, Michael D Altman, Lixia Li, Kumiko Nagashima, George N Naumov, Sriram Sathyanarayanan, Erica Cook, Erick Morris, Marc Ferrer, Bill Arthur, Yair Benita, Jim Watters, Andrey Loboda, Jeff Hermes, D Gary Gilliland, Michelle A Cleary, Pamela M Carroll, Peter Strack, Matt Tudor, Jannik N Andersen
The RAS-MAPK pathway controls many cellular programs, including cell proliferation, differentiation, and apoptosis. In colorectal cancers, recurrent mutations in this pathway often lead to increased cell signaling that may contribute to the development of neoplasms, thereby making this pathway attractive for therapeutic intervention. To this end, we developed a 26-member gene signature of RAS-MAPK pathway activity utilizing the Affymetrix QuantiGene Plex 2.0 reagent system and performed both primary and confirmatory gene expression-based high-throughput screens (GE-HTSs) using KRAS mutant colon cancer cells (SW837) and leveraging a highly annotated chemical library...
October 2016: Journal of Biomolecular Screening
https://www.readbyqxmd.com/read/27412535/development-of-a-3d-tissue-culture-based-high-content-screening-platform-that-uses-phenotypic-profiling-to-discriminate-selective-inhibitors-of-receptor-tyrosine-kinases
#18
Tijmen H Booij, Maarten J D Klop, Kuan Yan, Csaba Szántai-Kis, Balint Szokol, Laszlo Orfi, Bob van de Water, Gyorgy Keri, Leo S Price
3D tissue cultures provide a more physiologically relevant context for the screening of compounds, compared with 2D cell cultures. Cells cultured in 3D hydrogels also show complex phenotypes, increasing the scope for phenotypic profiling. Here we describe a high-content screening platform that uses invasive human prostate cancer cells cultured in 3D in standard 384-well assay plates to study the activity of potential therapeutic small molecules and antibody biologics. Image analysis tools were developed to process 3D image data to measure over 800 phenotypic parameters...
October 2016: Journal of Biomolecular Screening
https://www.readbyqxmd.com/read/27358388/identification-of-novel-inhibitors-of-the-type-i-interferon-induction-pathway-using-cell-based-high-throughput-screening
#19
Zoe O Gage, Andri Vasou, David W Gray, Richard E Randall, Catherine S Adamson
Production of type I interferon (IFN) is an essential component of the innate immune response against invading pathogens. However, its production must be tightly regulated to avoid harmful effects. Compounds that modulate the IFN response are potentially valuable for a variety of applications due to IFN's beneficial and detrimental roles. We developed and executed a cell-based high-throughput screen (HTS) targeting components that participate in and/or regulate the IRF3 and nuclear factor (NF)-κB branches of the IFN induction pathway...
October 2016: Journal of Biomolecular Screening
https://www.readbyqxmd.com/read/27256155/a-novel-automated-high-content-analysis-workflow-capturing-cell-population-dynamics-from-induced-pluripotent-stem-cell-live-imaging-data
#20
Maximilian Kerz, Amos Folarin, Ruta Meleckyte, Fiona M Watt, Richard J Dobson, Davide Danovi
Most image analysis pipelines rely on multiple channels per image with subcellular reference points for cell segmentation. Single-channel phase-contrast images are often problematic, especially for cells with unfavorable morphology, such as induced pluripotent stem cells (iPSCs). Live imaging poses a further challenge, because of the introduction of the dimension of time. Evaluations cannot be easily integrated with other biological data sets including analysis of endpoint images. Here, we present a workflow that incorporates a novel CellProfiler-based image analysis pipeline enabling segmentation of single-channel images with a robust R-based software solution to reduce the dimension of time to a single data point...
October 2016: Journal of Biomolecular Screening
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