Read "A simple high throughput assay to evaluate water consumption in the fruit fly, Scientific Reports" on DeepDyve, the largest online rental service for scholarly research with thousands of academic publications available at your fingertips.
Dr. Linda Baum and CDMD colleagues published a paper on a high throughput screen to find known drugs that might increase sugars on muscle cells as a possible new therapeutic intervention in DMD. Click for article ...
High‐content flow (HCF) screening systems, such as the iQue Screener and HTFC Screening System from IntelliCyt, have facilitated the implementation of flow cytometry assays for high‐throughput screening
Type: Heart. Grant holder: Dr Paul Welsh, Dr Preiss, Professors Woodward, Chalmers and Sattar; Cardiovascular Research Centre, University of Glasgow. Amount Awarded: £77,630 over two years. Year: 2013 - Finished. Cutting-edge technology is being harnessed to give a more detailed analysis of the chemical make-up of blood samples than has yet been possible. The research team has developed the ability to measure the levels of more than 100 different components in blood, covering a broad range of fats and sugars. The researchers will test the new technologys ability to predict heart attacks, thereby improving drug prescription and prevention. This new technology is being applied in a major international trial of diabetes patients, although this will be of particular interest in Scotland where rising diabetes rates threaten to reverse current declining heart attack trends.. ...
High-throughput cell-based screens have become an important experimental tool for the analysis of many cellular processes. Whole genome sequences and methods for gene silencing by RNA interference (RNAi) have enabled loss-of-function analysis in ex vivo and in vivo, opening new avenues for functional analysis that were previously unfeasible [1, 2]. Different experimental methods to assess phenotypic changes are being used, from single-channel homogenous readouts to multi-channel cytometry and imaging, producing large data sets that need to be analyzed to extract phenotypically relevant information. RNAi screening has found a broad user-base as a genetic method to dissect many different cellular processes, such as cell survival, signaling pathways and other cellular phenotypes in a high-throughput manner [3-6].. High-throughput screens are mostly performed using 96- to 384-well plates and produce large data sets that need to be normalized, summarized and ranked to generate a list of significant ...
Cytotoxicity tests for high-throughput drug discovery.: Despite theoretical obstacles associated with performing cell-based assays in high-density formats (micr
Included among the quantitative high throughput screens (qHTS) conducted in support of the US Tox21 program are those being evaluated for the detection of genotoxic compounds. One such screen is based on the induction of increased cytotoxicity in seven isogenic chicken DT40 cell lines deficient in DNA repair pathways compared to the parental DNA repair-proficient cell line. To characterize the utility of this approach for detecting genotoxic compounds and identifying the type(s) of DNA damage induced, we evaluated nine of 42 compounds identified as positive for differential cytotoxicity in qHTS (actinomycin D, adriamycin, alachlor, benzotrichloride, diglycidyl resorcinol ether, lovastatin, melphalan, trans-1,4-dichloro-2-butene, tris(2,3-epoxypropyl)isocyanurate) and one non-cytotoxic genotoxic compound (2-aminothiamine) for (1) clastogenicity in mutant and wild-type cells; (2) the comparative induction of γH2AX positive foci by melphalan; (3) the extent to which a 72-hr exposure duration ...
DasGupta et al. [3] developed a high-throughput assay based on the known ability of canonical Wnt signaling to activate transcription of luciferase reporter constructs in transfected cells. Improving on the widely used construct TOP-Flash [13], they generated two new reporters each containing multiple TCF-binding sites upstream of a different minimal promoter. Because only the TCF sites were common between the reporters, off-target effects unrelated to β-catenin/TCF signaling were minimized. Reporters with mutated TCF-binding sites also served as specificity controls. The authors first validated the behavior of these reporters in transfection assays of Drosophila cell lines. Then they scaled up the transfections to incorporate approximately 22,000 double-stranded RNAs (dsRNAs), so as to induce RNAi [3], and tested the individual effects on Wingless-induced signaling. The library of dsRNA sequences, previously used in other high-throughput RNAi screens, is directed at all known open reading ...
Understanding the tens of thousands of proteins that compose the ...The bulk of these so far enigmatic proteins may now be open to study ...The scientists have developed the first widely applicable high-through...Most studies of enzymes rely on substrate assays which allow research...One major hurdle in protein research though is that developing conve...,New,high-throughput,screening,technique,makes,probing,puzzling,proteins,possible,biological,biology news articles,biology news today,latest biology news,current biology news,biology newsletters
TY - JOUR. T1 - Application of a high throughput method of biomarker discovery to improvement of the EarlyCDT®-Lung Test. AU - Macdonald, Isabel K.. AU - Murray, Andrea. AU - Healey, Graham F.. AU - Parsy-Kowalska, Celine B.. AU - Allen, Jared. AU - McElveen, Jane. AU - Robertson, Chris. AU - Sewell, Herbert F.. AU - Chapman, Caroline J.. AU - Robertson, John F.R.. PY - 2012/12/13. Y1 - 2012/12/13. N2 - The National Lung Screening Trial showed that CT screening for lung cancer led to a 20% reduction in mortality. However, CT screening has a number of disadvantages including low specificity. A validated autoantibody assay is available commercially (EarlyCDT®-Lung) to aid in the early detection of lung cancer and risk stratification in patients with pulmonary nodules detected by CT.Recent advances in high throughput (HTP) cloning and expression methods have been developed into a discovery pipeline to identify biomarkers that detect autoantibodies. The aim of this study was to demonstrate the ...
in Journal of pharmacy & pharmaceutical sciences : a publication of the Canadian Society for Pharmaceutical Sciences, Societe canadienne des sciences pharmaceutiques (2013), 16(2), 217-30. PURPOSE: ATPases, which constitute a major category of ion transporters in the human body, have a variety of significant biological and pathological roles. However, the lack of high throughput assays for ... [more ▼]. PURPOSE: ATPases, which constitute a major category of ion transporters in the human body, have a variety of significant biological and pathological roles. However, the lack of high throughput assays for ATPases has significantly limited drug discovery in this area. We have recently found that the genetic deletion of the ATP dependent calcium pump PMCA4 (plasma membrane calcium/calmodulin dependent ATPase, isoform 4) results in infertility in male mice due to a selective defect in sperm motility. In addition, recent discoveries in humans have indicated that a single nucleotide polymorphism ...
Quantitative high resolution melting: two methods to determine SNP allele frequencies from pooled samples. . Biblioteca virtual para leer y descargar libros, documentos, trabajos y tesis universitarias en PDF. Material universiario, documentación y tareas realizadas por universitarios en nuestra biblioteca. Para descargar gratis y para leer online.
Gene-expression analysis is ubiquitously used both in life sciences and in medical research [1-3]. Microarray and reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) analyses are commonly used to measure transcript abundance. Microarrays allow the massive parallel analysis of thousands of genes but still require significant time and financial expenses. RT-qPCR is used as a conventional routine method for expression profiling of a moderate number of genes and is highly suitable when only a small amount of sample is available [1, 4]. High throughput thermal cyclers and PCR platforms help to overcome the limitation of RT-qPCR and to perform simultaneous expression analysis of tens and hundreds of genes for one or more samples depending on the platform [1].. In research practice these two technologies are able to solve a broad spectrum of questions supplementing each other. In clinical diagnostics RT-qPCR is still more popular with the advantage of speed and a relatively ...
Gentaur molecular products has all kinds of products like :search , MarkerGene \ MarkerGene™ Multiple Drug Resistance Microtiterplate Assay Kit, High-throughput assay system based on measurement of efflux of green fluorescent dye known to bind to MDR transporters ABCG2 and ABCB1 \ M1580 for more molecular products just contact us
EN) - High-throughput assay setup for the Investigator 24plex GO! Kit from Bode Buccal 2 Assembled Cassette samples using the STAR Q Punch ...
Use NucleoSpin 8/96 Blood QuickPure kits to isolate genomic DNA from whole blood, serum, plasma, and body fluids in flexible 8-well strip or high-throughput 96-well plate formats. Use animal or human blood samples, fresh or frozen.
Radiotherapy (RT) is one of the most commonly used treatments for cancer. Approximately 50% of all cancer patients are treated with RT. For many indications, radiotherapy is combined with other treatment modalities, such as surgery and/or chemotherapy [1-4]. The biological basis for the therapeutic effects of RT is that the applied ionizing radiation (IR) causes lethal double-strand breaks in the cellular DNA leading to tumor cell death. However, IR-induced DNA damage also triggers DNA damage response (DDR) signaling pathways in cells. These can result either in cell cycle arrest and DNA damage repair or in cell death. Differences in the functioning of these processes in different cells or under different conditions determine the final effect of a certain dose of IR [5]. Cancer cells are generally more vulnerable to DNA damage than healthy cells [6].. Despite its broad use and implementation of improved methods, clinical success of radiotherapy is variable. While survival rates after RT are high ...
To assess the quantitative ability of this approach, we performed a second experiment where we mixed fully methylated and nonmethylated DNA (the mixtures contained 0, 1, 5, 10, 20, 30, 40, 50, 60, 70, 80, 90, 95, 99, and 100% of methylated DNA) and estimated the corresponding relative methylation (see Fig. 3b ). The means and standard deviations shown are derived from six replicates of the PCR, transcription, cleavage, and MS measurement steps. In these mixing experiments we found a direct correlation between the estimated relative methylation based on the signal intensities of the two cleavage products and the relative amount of methylated template DNA. As depicted in Fig. 3b , the linear relationship between the actual and estimated relative methylation spans the entire range in these mixtures. The average standard deviation of relative methylation was ≈5% for mixtures between 10% and 90%. Outside of this range, quantitative data can be analyzed, but limitations in the dynamic range of ...
In this study, we identified a novel class of CRAC channel blockers using high-throughput chemical library screen and asked how inhibition of Ca2+ signaling during TCR stimulation influenced differentiation and effector functions of T cells in vitro and in vivo. Using a combination of NFAT translocation as readout and a cell line harboring amplified CRAC currents, we identified a novel class of immunomodulators, compound 5, and its analogs. A more potent analog of the lead compound, compound 5D, blocked CRAC currents generated by WT Orai1 and a constitutively active mutant of Orai1 without affecting Orai1 and STIM1 translocation, indicating that the blocking mechanism directly involves the pore-forming subunit, Orai1. Further studies of CRAC current inhibition showed block by extracellular, but not intracellular, application of compound 5D, suggesting that the binding site is accessible only from the extracellular face of Orai1. Comprehensive mutational analysis of all the residues with possible ...
The process of washing suspension cells - pipetting, centrifuging, and dispensing - can be very disruptive and lead to loss of reagents and cells and cause unnecessary strain. In addition, the procedure is overly time consuming and is subject to many error-prone steps. Overall, these limitations make it unfeasible to perform many cellular assays with suspension cells, including high throughput assays.. The DropArray platform lets these cells be retained, allowing entire assays to be performed directly on the proprietary, wall-less microtiter plate - the DropArray Microplate. Its unique design allows researchers to culture as little as 250 cells (384-format) or 750 cells (96-well format) or as many as 10,000 cells (384-well format) or 40,000 cells (96-well format) per assay. In addition, plate washing is performed using gentle, low-velocity liquid exchange that keeps suspension cells within the well. Overall, this enables fully automatable, plate-based assays with suspension cells that were ...
PubMed comprises more than 30 million citations for biomedical literature from MEDLINE, life science journals, and online books. Citations may include links to full-text content from PubMed Central and publisher web sites.
We have developed a novel high-throughput screening platform for the discovery of small-molecules catalysts for bond-forming reactions. The method employs an in vitro selection for bond-formation using amphiphilic DNA-encoded small molecules charged with reaction substrate, which enables selections to be con
Computational and statistical methods development for high-throughput, high-content biological data. Modeling and integration of data from high throughput assays for biomarker discovery, clinical outcome prediction and disease classification.
From the brief descriptions of the project I have been able to find, the proposed NIH research plan will only deal with mutations in single genes. Unless the "high-throughput" analyses that go unnamed consist of karyotyping or in situ probes for duplications, this study wont even skim the surface of the genetics of cancer. I dont think you can even do either of these in a high-throughput manner. The epigenetic effects may be picked up by the gene expression analysis, but it will be difficult to distinguish between mutations to regulatory regions (cis effects), mutations to transcription factors that control the expression of the gene (trans effects), and epigenetic effects.. Without more detail, its hard to judge the merits of this project. The research will definitely result in some worthwhile discoveries, but it may be wiser to spend the money on more detailed analyses. Just because "high-throughput" works for genome sequencing, doesnt mean that Francis Collins needs to apply it to every ...
Skaga, Erlend; Kulesskiy, Evgeny; Brynjulfsen, Marit; Sandberg, Cecilie Jonsgar; Kyttala, Aija; Langmoen, Iver Arne; Laakso, Aki; Gaal-Paavola, Em-Lia; Perola, Markus; Wennerberg, Krister; Vik-Mo, Einar ...
A critical unmet need in implementing personalized medicine is the ability to sort through the millions of single nucleotide polymorphisms (SNPs) present in the human genome and to pinpoint which of these DNA variations are causative in disease. A key under-studied function of SNPs is their ability to generate or disrupt genomic binding sites for transcription factors involved in cancer. Toward this goal, we are inventing the SNP-SNAP (Specificity and Affinity for Proteins) microarray as a prototype high throughput device to evaluate SNP function. The SNP-SNAP arrays will be used to display a quarter-million prostate cancer- related SNPs as double-stranded DNA molecules and to assay transcription factors (i.e., drug targets) for their binding to these SNP DNA sequences. The resulting data will be correlated with prostate cancer incidence. The million-plus data points from the SNP-SNAP arrays will be analyzed using SNP-Sequence Specificity Landscapes, creating a prostate cancer molecular ...
In this study, we present a quantitative high-throughput assay with reduced costs for methylation analysis. On the MALDI-TOF MS, mass spectrum information can be used to determine methylated and unmethylated DNAs to assess the degree of methylation for each CpG island independently, and to estimate the average methylation for the entire target region (5, 6, 13, 14). Using this method, both hypermethylation and hypomethylation can be detected in samples.. Recently, the company developed a product that showed robustness of (C/T) cleavage with the MassARRAY system for methylation analysis based on a study assessing a set of ,400 candidate genes in 59 different cancer cell lines (6). In the MassCLEAVE kit (Sequenom, Inc.), the T-cleavage reagent contains dCTP, rUTP, rGTP, and rATP, and the C-cleavage reagent contains a mixture of dTTP, rCTP, rGTP, and rATP, resulting in a unique cleavage 3′ of rCTP or rUTP only, respectively. In EpiTYPER two cleavage reactions, the reverse strand is cleaved by ...
Video, Wyss researchers have created a high-throughput platform to generate an Adeno-associated virus 2 (AAV2) library containing 200,000 variants, each carrying a distinct mutation in the virus capsid protein.
The voltage-sensitive L-type Ca2+-channel (LTCC) CaV1.2 is a crucial component for controlling intracellular activity and thereby essential in the cardiovascular and neuronal system. It is widely expressed in vascular smooth muscle tissue and the heart muscle 1-3. The opening of the channels leads to an increase of intracellular calcium, which act as second messenger, and thereby affects a variety of cellular processes 4 including heart muscle contraction and CaV1.2 is therefore an important target in e.g. safety pharmacology screening. CaV1.2 channels are known to require a large depolarization for their activation and once activated they display a long-lasting current flow, which typically can be blocked by low micromolar concentrations of e.g. dihydropyridines, phenylalkylamines and benzothiazepines 5,6.. In these studies, currents from HEK-hCaV1.2 were recorded on the high-throughput platform Qube 384 in both single-hole and multi-hole mode. Success rates, IV characteristics and the ...
Love J, Mancia F, Shapiro L, Punta M, Rost B, Girvin M, Wang D-N, Zhou M, Hunt JF, Szyperski T, et al. The New York Consortium on Membrane Protein Structure (NYCOMPS): a high-throughput platform for structural genomics of integral membrane proteins. J Struct Funct Genomics. 2010 ;11(3):191-9. ...
Love J, Mancia F, Shapiro L, Punta M, Rost B, Girvin M, Wang D-N, Zhou M, Hunt JF, Szyperski T, et al. The New York Consortium on Membrane Protein Structure (NYCOMPS): a high-throughput platform for structural genomics of integral membrane proteins. J Struct Funct Genomics. 2010 ;11(3):191-9. ...
Hepatitis C virus (HCV) infection affects an estimated 185 million people worldwide, with chronic infection often leading to liver cirrhosis and hepatocellular carcinoma. Although HCV is curable, there is an unmet need for the development of effective and affordable treatment options. Through a cell-based high-throughput screen, we identified chlorcyclizine HCl (CCZ), an over-the-counter drug for allergy symptoms, as a potent inhibitor of HCV infection. CCZ inhibited HCV infection in human hepatoma cells and primary human hepatocytes. The mode of action of CCZ is mediated by inhibiting an early stage of HCV infection, probably targeting viral entry into host cells. The in vitro antiviral effect of CCZ was synergistic with other anti-HCV drugs, including ribavirin, interferon-α, telaprevir, boceprevir, sofosbuvir, daclatasvir, and cyclosporin A, without significant cytotoxicity, suggesting its potential in combination therapy of hepatitis C. In the mouse pharmacokinetic model, CCZ showed ...
Kinome microarrays are comprised of peptides that act as phosphorylation targets for protein kinases. This platform is growing in popularity due to its ability to measure phosphorylation-mediated cellular signaling in a high-throughput manner. While software for analyzing data from DNA microarrays has also been used for kinome arrays, differences between the two technologies and associated biologies previously led us to develop Platform for Intelligent, Integrated Kinome Analysis (PIIKA), a software tool customized for the analysis of data from kinome arrays. Here, we report the development of PIIKA 2, a significantly improved version with new features and improvements in the areas of clustering, statistical analysis, and data visualization. Among other additions to the original PIIKA, PIIKA 2 now allows the user to: evaluate statistically how well groups of samples cluster together; identify sets of peptides that have consistent phosphorylation patterns among groups of samples; perform hierarchical
The new version includes: Astrocytes lineage ; Updates to Kidney, Pancreas, Bone and Cartilage (from somite & neural crest) lineages ; 19 new high-throughput experiments for: hair, tooth, early embryo, cornea, lens and astrocytes ; New data from Bodymap & RNAseq ; 40 new protocols, including 7 new categories of direct reprogramming protocols added ; ~60 new patient-derived iPSCs ; Family cell descriptions for BM-MSCs, Adipose-derived MSCs and UC-MSCs (tissue) and 50 new cell therapies, including 7 cards of marketed cell-based products. Additionally, The UCB-MSCs (blood) cell family was split into tissue and cord blood with full elaborate descriptions. ...
GENEWIZ Implements Edico Genome DRAGEN Processors to Accelerate Analysis of Big Data Generated by New High-Throughput, Next-Generation Sequencing Platforms
As noted above, the MTT assay is really a metabolic assay because the MTT molecule needs to enter a cell and get converted to Formazan using NADPH. While the exact mechanism of MTTs metabolism isnt clear, this means the mitochondria needs to be intact and functioning. So, if you add a cytotoxic material which reduces mitochondrial efficiency, you might get weird results. In this case, its useful to also know other live/dead assays. The other major cell viability assays that are used in research include:. Cell Titer Blue: Similar to the MTT Assay, this assay involves incubating cells with resazurin (blue) and forming resorfurin (pink) after the cells metabolize it. Generally the metabolism takes 1-4 hours but it is much more sensitive than the MTT assay because you can measure the product via fluorescence (Ex/Em 560 nm/590 nm). The main advantage of this assay is that you dont need to resolubilize the product in DMF/SDS so its much simpler. This is also a great high throughput assay!. Trypan ...
A body 300 having a cavity 310 for mounting a substrate 120 fabricated with probe sequences at known locations according to the methods disclosed in U.S. Pat. No. 5,143,854 and PCT WO 92/10092 or others, is provided. The cavity includes inlets 350 and 360 for introducing selected fluids into the cavity to contact the probes. Accordingly, a commercially feasible device for use in high throughput assay systems is provided.
A body 300 having a cavity 310 for mounting a substrate 120 fabricated with probe sequences at known locations according to the methods disclosed in U.S. Pat. No. 5,143,854 and PCT WO 92/10092 or others, is provided. The cavity includes inlets 350 and 360 for introducing selected fluids into the cavity to contact the probes. Accordingly, a commercially feasible device for use in high throughput assay systems is provided.
Cellular kinases play an important role in relaying signals from activated receptors residing at the cell membrane to the interior of the cell, through signal transduction. The cellular processes in which they are associated include angiogenesis, cell growth, cell migration, and apoptosis. In this application note from BioTek, a method that uses HTRF® technology to quantify cellular kinase activity of VEGFR2 and STAT3 without overexpression of kinase or its substrate is described. Automation of
Methods for determining a function of cells, which comprises a suspension of cells flowing along a first fluid channel. The cells have a first detectable property associated therewith, and wherein the cells produce a second detectable property upon activation of the function of the cells, the first and second detectable properties being distinguishable from each other. Levels of the first and second detectable properties are measured. The level of the second detectable property is compared to the level of first detectable property to determine the relative function of the cells.
Most of the work in this area has focused on mapping protein-protein interactions (e.g., see Stanyon and Finley, 2000; Zhong et al., 2003; Giot, 2003; Stanyon et al., 2004; Parrish et al., 2007). A long running project has been to construct and characterize a proteome-wide interaction map for Drosophila. This project entails conducting high throughput screens, developing computational tools to analyze interaction data, and sharing the interaction data and analysis tools with the public via a web-accessible database (www.DroIDb.org). We have also been concerned with figuring out how to use the flood of functional genomic data from our own and other studies to create systems-wide models of biological processes. In addition to the public database and network analysis interfaces (Murali et al., 2011; Pacifico et al., 2006; Yu et al., 2008), the lab has developed computational methods to analyze interaction networks (Murali et al., 2014), to predict new protein interactions (Yu et al., 2005), to ...
We have received encouraging biological results for the analogues we sent for testing before Christmas. Mat has discussed this here on TSL and on G+. Our best hit came from the "near-neighbour" compound and the original GSK hits came out slightly less active than in their original high throughput screen. However, Paul Willis at MMV rates TCMDC-123794 as a better lead than PMY 14-1 (TSL post). Ive already started on the synthesis of the amide (PMY 31) and ether-linked analogues as well as the "linker-less" analogues (PMY 12-5). As mentioned by Paul, it could well be that inherent instability of the ester link would diminish the activity of the compound. We have already found that the free acid and methyl esters themselves are inactive so inclusion of amide or ether linkers could increase activity or at least prolong the metabolic half-life ...
We have received encouraging biological results for the analogues we sent for testing before Christmas. Mat has discussed this here on TSL and on G+. Our best hit came from the "near-neighbour" compound and the original GSK hits came out slightly less active than in their original high throughput screen. ...
To measure mutation frequency in the recovered plasmid, plasmid DNA sequence was determined using an M13 (−47) primer. Use long extensiontimes (at least 3 min).If this fails, test the primers, template andother reagentsunder "normal" PCR conditions to ensurethat there has notbeen a primer design or synthesis error, ora degeneration Next, you could transform the library into a strain where the receptor would be expressed and apply a high throughput screen to pick out variants in the library that have the Schematic diagram of error-prone RCA in comparison with the conventional random mutagenesis methods. For more information see here and here (see page 13). The fraction of un-mutated DNA templates as a function of template length and number of EP-PCR doublings.EP-PCRdoublingsmutations per nucleotide position100 bp200 bp400 bp800 bp1600 ...
Methods of producing liquid handling biosensor devices are provided. The liquid handling biosensor devices allow detection of biomolecular interactions in liquid. The use of labels is not required and the methods can be performed in a high-throughput manner.
Quantifying DNA-protein interaction using DNA microarrays are gaining increasing attention due to their ability to profile specificity of interactions in a high-throughput manner. This paper describes a new approach that used the ability of ssDNA-dsDNA probe to complex with DNA binding proteins in the solution phase and then spatially immobilized onto microarray through specific DNA hybridization. In one case, the Spatially Addressable DNA Array (SADA) approach demonstrated that enzymatic cleavage in solution is more efficient than if conducted heterogeneously. In addition, binding of RNA polymerase with promoter DNA could be detected with this strategy ...
The present invention provides a technology called Pulse-Multiline Excitation or PME. This technology provides a novel approach to fluorescence detection with application for high-throughput identification of informative SNPs, which could lead to more accurate diagnosis of inherited disease, better prognosis of risk susceptibilities, or identification of sporadic mutations. The PME technology has two main advantages that significantly increase fluorescence sensitivity: (1) optimal excitation of all fluorophores in the genomic assay and (2) "color-blind" detection, which collects considerably more light than standard wavelength resolved detection. Successful implementation of the PME technology will have broad application for routine usage in clinical diagnostics, forensics, and general sequencing methodologies and will have the capability, flexibility, and portability of targeted sequence variation assays for a large majority of the population ...
Ralph Garippa, Ph.D., independent consultant and former head of cell-based high-throughput screening (HTS) and microscopic imaging-based high-content
Our goal is to investigate the utility of cancer molecular phylogeny, a novel technological concept, to predict clinical outcomes. Rather than use novel high-throughput technologies to identify a molecular marker or signature to predict survival, we propose to use a molecular measure of cancer age. Estimation of cancer age is complicated by the fact that we do not observe the initial transformation and clonal expansion. What we do observe is a population of cells that are descendants of the original transformed cell. What we can measure is the molecular diversity of the population. Population genetics dictates that cells from an older population will show more diversity than cells from a younger population. Thus a molecular measure of tumor diversity should capture tumor age. We propose that tumor age will help predict patient outcomes. Thus, we challenge the current paradigm of finding a common pathway of cancer development that leads to poor survival. Instead we propose that older tumors are ...
Messenger RNA (mRNA) export adaptors play an important role in the transport of mRNA from the nucleus to the cytoplasm. that the histone chaperone FACT specifically binds UIF VE-821 but not REF via the SSRP1 subunit and this interaction is required for recruitment of UIF to mRNA. Together the results indicate that REF and UIF represent key human adaptors for the export of cellular mRNAs via the UAP56-NXF1 pathway. and proteins. Figure?1 Identification and Characterization of UIF By using an antiserum raised to UIF we detected it in extracts from 293T cells as a 37 kDa protein (Figure?1D). The levels of UIF were increased when cells were transfected with a UIF cDNA expression vector and reduced when cells expressed a microRNA (miRNA) targeting UIF messenger RNA (mRNA) indicating that the antibody recognized UIF. We analyzed manifestation of UIF in chick embryos and discovered a widespread manifestation pattern during advancement (Shape?S2). The expression pattern was identical compared to that ...
Culture-independent high-throughput sequencing has provided unprecedented insights into microbial ecology, particularly for Earths most ubiquitous and diverse inhabitants - the viruses. A plethora of methods now exist for amplifying the vanishingly small amounts of nucleic acids in natural viral communities in order to sequence them, and sequencing depth is now so great that viral genomes can be detected and assembled even amid large concentrations of non-viral DNA. Complementing these advances in amplification and sequencing is the ability to physically link fluorescently labeled viruses to their host cells via high-throughput flow sorting. Sequencing of such isolated virus-host pairs facilitates cultivation-independent exploration of the natural host range of viruses. Within the next decade, as these technologies become widespread, we can expect to see a systematic expansion of our knowledge of viruses and their hosts. | Viruses, Immunology & Bioinformatics from Virology.uvic.ca
Fueled by the fast-growing DNA sequence information, proteomics-the large-scale analysis of proteins-has become one of the most important disciplines to characterize protein activities and provide insight into functional network between protein molecules in a high-throughput format. More and more evidence has pointed out that proteins rarely act as single isolated species when performing their functions in vivo; they normally associate with other proteins and/or molecules as complexes and function in a network mode. The goal of my laboratory is to discover and characterize the activities of large collection of proteins or an entire proteome as the first step in better understanding the mechanisms of biological processes. More specifically, the lab is interested in analyzing protein posttranslational modifications, various signaling network, protein profiling, and host-pathogen interactions on the proteomics level. It is obvious that this type of research requires high-throughput technologies ...
Berkeley Lab researchers including Yasuo Yoshikuni, Gaoyan Wang, Zhiying Zhao, and Jan-Fang Cheng have developed a strategy to chromosomally integrate and express DNA constructs comprising single genes to complex pathways in a broad range of bacterial hosts. By comparing traditional product and pathway specific modifications to host strains in a high-throughput format, the Berkeley Lab technology reduces the time and cost of identifying candidate production strains in a significant way. Strain-to-strain variation can be minimized due to the use of a pre-defined chromosomal integration site. This development is a major technical step forward in the wholesale transfer of heterologous biosynthetic pathways to bacterial organisms derived from various environments across human, livestock, and plant microbiomes. The technology has been demonstrated to work across a wide variety of bacterial hosts. The Berkeley Lab technology allows for rapid and efficient transfer and integration of biosynthetic ...
We have developed spotted cell microarrays for measuring cellular phenotypes on a large scale. Collections of cells are printed, stained for subcellular features, then imaged via automated, high-throughput microscopy, allowing systematic phenotypic characterization. We used this technology to identify genes involved in the response of yeast to mating pheromone. Besides morphology assays, cell microarrays should be valuable for high-throughput in situ hybridization and immunoassays, enabling new classes of genetic assays based on cell imaging ...
New microscopy techniques are continuously developed, resulting in more rapid acquisition of large amounts of data. Manual analysis of such data is extremely time-consuming and many features are difficult to quantify without the aid of a computer. But with automated image analysis biologists can extract quantitative measurements and increases throughput significantly, which becomes particularly important in high-throughput screening (HTS). This thesis addresses automation of traditional analysis of cell data as well as automation of both image capture and analysis in zebrafish high-throughput screening. It is common in microscopy images to stain the nuclei in the cells, and to label the DNA and proteins in different ways. Padlock-probing and proximity ligation are highly specific detection methods that produce point-like signals within the cells. Accurate signal detection and segmentation is often a key step in analysis of these types of images. Cells in a sample will always show some degree of ...
By exploiting high-throughput microscopy and a new cell line, researchers have uncovered how the transcription regulator estrogen receptor-α can
The measurement of -L2AX foci induction in cells provides a sensitive and reliable method for the quantitation of DNA harm responses in a variety of cell types. in the DNA-PKcs defective cells (XP14BRneo17), we noticed an more advanced preservation of foci in the nuclei a sign of incomplete fix of DNA DSB. In overview, the program of image resolution stream cytometry provides allowed an evaluation of foci in a huge amount of cells (20,000) for each cell series at each period stage. This provides a story technique to determine distinctions in fix kinetics between different cell types. We recommend that image resolution stream cytometry provides an choice system for accurate computerized high through-put evaluation of foci induction in a range of cell types. ? 2011 Cosmopolitan Culture for Advancement Troxacitabine of Cytometry gene which features in the control of cell routine criminal arrest and induction of DNA DSB fix outcomes in the tenacity of -L2AX foci in the nucleus of cells shown to IR ...
Type 1 diabetes is associated with T\cell responses to \cell antigens such as GAD65. a potentially autoreactive repertoire. Without depleting CD25+ cells, GAD113C132 and GAD265C284 responses were significantly stronger in subjects with diabetes. Although nearly every individual responded to at least one GAD65 epitope, most were seen in less than half of the subjects tested, suggesting that multiple epitopes are recommended for TSPAN9 immune monitoring. biotinylation, class II monomers were loaded with either peptide pools or individual peptides by incubating for 48?hr at 37 with 25\fold molar excess of peptide (total) in phosphate buffer, pH 60 in the presence of 02% culture growth using CD25 microbeads (Miltenyi Biotec) as previously described to remove regulatory T cells and increase the magnitude of responses.19 In a second set of experiments, responses were evaluated without removing CD25+ cells. CD4+ T cells (or CD4+?CD25 T cells) were seeded in 48\well plates at 25??106?cells/well in ...
By the end of 2011 we will likely know the DNA sequences for 30,000 human genomes. However, to truly understand how the variation between these genomes affect phenotype at a molecular level, future research projects need to analyze these genomes in conjunction with data from multiple ultra-high throughput assays obtained from large sample populations. In cancer research, for example, studies that examine 1000s of specific tumors in 1000s of patients are needed to fully characterize the more than 10,000 types and subtypes of cancer and develop diagnostic biomarkers. These studies will use high throughput DNA sequencing to characterize tumor genomes and their transcriptomes. Sequencing results will be validated with non-sequencing technologies and putative biomarkers will be examined in large populations using rapid targeted assay approaches ...
Chromatin structure, genome architecture, epigenetics, hematopoiesis, stem cell biology, and cancer biology. RESEARCH DESCRIPTION. The focus of our research is on understanding the mechanisms underlying development and tumorigenesis through deciphering the structure-function relationship of mammalian genomes. We have developed a series of high throughput methods for mapping three-dimensional (3D) genome architecture globally or locally. Recently, in collaboration with the investigators in the University of Washington Center for Nuclear Organization and Function (UW-CNOF), we developed a single-cell Hi-C method for characterize the 3D genome architecture in thousands of single cells in a massively parallel fashion. We have used these tools to dissect the dynamics of 3D genome organization in the context of a variety of biological systems. One of our current focuses is to characterize the nuclear morphology-associated 3D genome rearrangement and epigenetic programming during blood cell maturation ...
Screening vs. Confirmation Low cost Fast Semi-quantitative High sensitivity Low specificity High cost Slow Quantitative High sensitivity High specificity
Course organised by LCBU and gives a practical basis for assay development in high throughput screening and is given as a practical complement to the course
High-content screening (HCS), also known as high-content analysis (HCA), is a method that is used in drug discovery to identify substances that alter...
Once a library of mutants is generated, they must be evaluated for their ability to perform the desired function. To do this, protein engineers employ a variety of high-throughput assays. Successful assays allow research to test a large number of functional variants while maintaining a connection between phenotype (the evolved protein function) and genotype (the DNA sequence encoding the evolved function). These assays can be categorized as either "in vivo" versus "in vitro" or "selection" versus "screening." The most important distinction is between a screen and a selection. Selections allow for only cells expressing proteins that exhibit the desired function to survive. In contrast, a screen allows for cells expressing any functional variant to survive yet be distinguished by phenotype. The most basic cell-based screening methods involve transforming a library of mutants into bacteria and identifying individual colonies or cultures that exhibit the desired function. These assays maintain a ...
High-throughput RNAi screening is widely applied in biological research, but remains expensive, infrastructure-intensive and conversion of many assays to HTS applications in microplate format is not feasible. Here, we describe the optimization
EDSP21計畫改以電腦模擬(In Silico)和高通量體外檢測(In Vitro High-Throughput Assays),加速對內分泌干擾物的篩檢
There have been many improvements in the area of cell counting and several instruments have been developed to move away from time-consuming manual cell counting. However, even with automated devices, the lack of throughput for cell concentration and viability measurements can create a process bottleneck, particularly at large-scale or industrial manufacturing. In addition, this lack of high-throughput can limit experimental design or create resource demand that isnt feasible when large numbers of samples need to be analyzed.
Optimization of a high-throughput 384 well virus replication assay.A. Comparison of virus replication over time (cells/viral foci) following infection using a s
Vol 30: HTML5 PivotViewer: high-throughput visualization and querying of image data on the web.. . Biblioteca virtual para leer y descargar libros, documentos, trabajos y tesis universitarias en PDF. Material universiario, documentación y tareas realizadas por universitarios en nuestra biblioteca. Para descargar gratis y para leer online.
The NucleoSpin 96 DNA RapidLyse kit allows rapid, high-throughput gDNA extraction from organs, cells, and tissues with the highest efficiency using a a unique lysis chemistry.
Visit the event Automated DNA Preparation and PCR Setup for High-Throughput Food and Feed Analysis at LABVOLUTION from 16 to 18 May. 2017 in Hannover, Germany. Plan your visit to the trade fair and search for further events.
The Namur Medicine & Drug Innovation Center (NAMEDIC) combines the expertise of the Chemistry and the Pharmacy Departments of the UNamur, with the aim to advance research on effective and innovative drugs. It is dedicated to medicinal chemistry, from hit identification to lead optimization.
A main presentation track at SLAS 2018 focuses on Assay Development and Screening with emphasis on methods such as high throughput screening, phenotypic screening, and 3rd and 4th dimension screens of cell-based assays.
Targeted gene- and protein-specific studies report protein regulation at the levels of protein synthesis or protein degradation (15, 45); yet, few global techniques have been developed to efficiently identify multiple targets of posttranscriptional regulation. Our developed methodology uses 2 standard high-throughput approaches tethered to computational analysis to systematically identify targets of posttranscriptional regulation. Our approach has the potential to identify a plethora of novel regulatory strategies because it can be applied to other perturbations and model systems. What we have demonstrated is that matched transcript-protein level studies can be used to identify discrepancies in standard gene regulatory models (i.e., transcript is induced to make more protein). The identification of any discrepancy or unusual regulatory pattern can be challenging to comprehend but detailed study ultimately leads to a better understanding of biology. Here we focused our studies on the large ...
Uncover deep biological understanding in your everyday assays and innovative applications using the Operetta CLS™ high-content analysis system. Featuring a unique combination of technologies, the system delivers all the speed, sensitivity and resolution you need to reveal fine sub-cellular details. And with our simple, powerful Harmony 4.5 software, Operetta CLS™ lets you find even subtle phenotypic changes.
In the searing mid-morning heat, Simon Peter Oola sweats profusely as he wheels three bags of rice into a large compound, where his produce must be weighed and certified before ...leer más ...
BioAssay record AID 488848 submitted by Broad Institute: Apoptosis Measured in Cell-Based System Using Plate Reader - 2061-02_Inhibitor_Dose_DryPowder_Activity.
Read user reviews, compare products and contact manufacturers of High Throughput Screening (HTS) products, including fluorescence, polarization and TR FRET readers, assays and robotic workstations on SelectScience.
The global High Throughput Screening Market is poised to reach USD 21.69 Billion in 2023. HTS Market Report provides crucial industry insights that will help your business grow.
High throughput protein expression to optimize your protein productions. 50x yield improvement / high-performance expression systems / 1000 tests in 4 weeks.
Introduction The amounts below are given for a 12-well plate format. Use Table 1 to adjust the reagent volumes for other plate sizes
... 具有很好的热稳定性,分散性,生物相容性,高的荧光稳定性和表面可修饰性。可提供多种不同粒径大小,不同颜色,不同荧光通量的荧光微球,可广泛应用与生物检测、疾病诊断、免疫检测、液相芯片技术、高通量药物筛选(HTS)等生物医学领域 ...
The technology provides numerous systems and methods to detect stress in stem cells by measuring potency factors (and markers) and prioritized differentiation factors (and markers). "Potency factors" can be thought of as "brake pedals" for differentiation. That is, their expression and activity is associated with toti- or pluri-potency. "Differentiation factors" can be thought of as "accelerator pedals" for differentiation. That is, their expression and activity is associated with normal differentiation that occurs when potency-maintaining extracellular growth factors are removed. Under stress conditions, however, potency factors can decrease and differentiation factors can increase even in the presence of potency-maintaining extracellular growth factors. This type of differentiation is referred to as "compensatory" or "prioritized" differentiation. That is, under compensatory or prioritized differentiation, stem cells can be induced by stress from a toti- or pluripotent state into a more ...
To screen for chemical toxicity in human or mouse stem cell lines (undifferentiated or differentiated) by quantitative high throughput screening (qHTS) at NCGC, or by using lower throughput assays at NIEHS. Initially, the project focuses on fostering collaborations with stem cell technology providers and assessing control compounds and subsets of NTP chemicals using various assay approaches. Stem cell technology platforms and model systems shown to be useful for in vitro toxicology screening will be employed with larger sets of chemicals for hazard identification and chemical prioritization for toxicity testing. Data have been generated on a library of 80 predominantly developmental neurotoxicants evaluated for effects on neurite outgrowth in a human stem cell-derived neural cell population, cytotoxicity in different neural populations derived from human stem cells, and effects on the beating of human stem cell derived cardiomyocytes. Dose-response analysis has been carried out on the data from ...
Drugs currently available for leishmaniasis treatment often show parasite resistance, highly toxic side effects and costs that are prohibitive commonly for patients from the tropical endemic countries. There is an urgent need for new drugs as a treatment solution for this neglected disease. Here the authors describe the development and implementation of an automated high-throughput viability screening assay for the discovery of new drugs against Leishmania. Assay validation was done with Leishmania promastigote forms, and included the screening of 4000 compounds with known pharmacological properties. In an attempt to find new compounds with leishmanicidal properties, 26,500 structurally diverse chemical compounds were screened. A cut-off of 70% growth inhibition in the primary screening led to the identification of 567 active compounds. Cellular toxicity and selectivity were responsible for the exclusion of 78% of the pre-selected compounds. The activity of the remaining 124 compounds was ...
Patients with hereditary diffuse gastric cancer (HDGC), a cancer predisposition syndrome associated with germline mutations of the CDH1 (E-cadherin) gene, have few effective treatment options. Despite marked differences in natural history, histopathology, and genetic profile to patients afflicted by sporadic gastric cancer, patients with HDGC receive, in large, identical systemic regimens. The lack of a robust preclinical in vitro system suitable for effective drug screening has been one of the obstacles to date which has hampered therapeutic advances in this rare disease. In order to identify therapeutic leads selective for the HDGC subtype of gastric cancer, we compared gene expression profiles and drug phenotype derived from an oncology library of 1912 compounds between gastric cancer cells established from a patient with metastatic HDGC harboring a c.1380delA CDH1 germline variant and sporadic gastric cancer cells. Unsupervised hierarchical cluster analysis shows select gene expression alterations
The WHO gold standard method for malaria diagnosis is microscopy. It is considered inexpensive and field adapted, even though it is time-consuming and requires specifically trained personnel. Molecular detection methods achieve much higher detection sensitivities [1, 26], and they are better adapted to automation of the process and objective reading of results by machines. This potential makes them a valuable option for large-scale epidemiologic studies. Unfortunately few efforts have been spent on developing the high-throughput approaches needed for such studies.. The 18S rRNA gene is the most frequently cited marker for malaria detection. It is composed of highly conserved regions which can be targeted for a qualitative detection of Plasmodium spp., and of variable zones allowing species identification [4-7]. However, the 18S rRNA genes in Plasmodium spp. also have unusual properties, such as the existence of three stage-specific A-, S- and O-types, as well as copy number and strain-specific ...
We present a novel automated methodology that compensates for the effect of cell morphology on flow cytometry data, and thereby enables a quantitative analysis of high‐throughput flow cytometry data. The algorithm normalizes the effect of the physical characteristics of cell size and cell granularity on the fluorescence intensity, thereby enabling the analysis of fluorescence intensities (protein abundance) in the presence of different morphological characteristics of cells in a population. In contrast to traditional gating, which discards the large majority of cells, the regression model retains all cells and thereby provides more accurate statistics, higher consistency across replicates and the ability to handle biological samples that contain far fewer cells (at least 10‐fold), allowing for faster and cheaper data acquisition. This is relevant when one is looking for rare cells (e.g., stem cells), or when performing high‐throughput screens where only a few hundred cells per experimental ...
To understand the function of cellular protein networks, spatial and temporal context is essential. Fluorescence correlation spectroscopy (FCS) is a single-molecule method to study the abundance, mobility and interactions of fluorescence-labeled biomolecules in living cells. However, manual acquisition and analysis procedures have restricted live-cell FCS to short-term experiments of a few proteins. Here, we present high-throughput (HT)-FCS, which automates screening and time-lapse acquisition of FCS data at specific subcellular locations and subsequent data analysis. We demonstrate its utility by studying the dynamics of 53 nuclear proteins. We made 60,000 measurements in 10,000 living human cells, to obtain biophysical parameters that allowed us to classify proteins according to their chromatin binding and complex formation. We also analyzed the cell-cycle-dependent dynamics of the mitotic kinase complex Aurora B/INCENP and showed how a rise in Aurora concentration triggers two-step complex ...
In collaboration with Dr. Lisa DeLouise, the Kobie Lab has developed patented nanowell array technology called microbubbles, which allow for the long term culture and functional interrogation of B cells at the single cell level. This high-throughput platform enables the rapid screening of millions of individual B cells to obtain precise resolution of cellular diversity and isolation of rare B cells with desired functional profiles. This platform is particularly advantageous for high-content single cell analysis and therapeutic monoclonal antibody development.. « back to all projects. ...
In contrast to upstream optimization, which focuses mainly on preservation of cells and proteins as they grow, downstream optimization focuses primarily on achieving maximum yield from each lot, and ensuring that the resulting product remains as concentrated and high-quality as possible. These improvements may be achieved by implementing additional filtration and purification steps, introducing new high-throughput methods, and/or re-engineering existing processes to streamline the pipeline from cell harvest to final product.. Chromatographic separation methods are often used to isolate mAbs (or other desired proteins) from the fermentation broth after harvesting. Chromatographic separation is highly selective, resulting in a significantly higher flow and purity rates than conventional filtration techniques. In fact, recent advances have helped further integrate chromatographic separation into the production process, allowing for fine-tuned adjustment of elution conditions, thus effectively ...
The virologic synapse (VS), which is formed between a virus-infected and uninfected cell, plays a central role in the transmission of certain viruses, such as HIV and HTLV-1. During VS formation, HTLV-1-infected T-cells polarize cellular and viral proteins toward the uninfected T-cell. This polarization resembles anterior-posterior cell polarity induced by immunological synapse (IS) formation, which is more extensively characterized than VS formation and occurs when a T-cell interacts with an antigen-presenting cell. One measure of cell polarity induced by both IS or VS formation is the repositioning of the microtubule organizing center (MTOC) relative to the contact point with the interacting cell. Here we describe an automated, high throughput system to score repositioning of the MTOC and thereby cell polarity establishment. The method rapidly and accurately calculates the angle between the MTOC and the IS for thousands of cells. We also show that the system can be adapted to score anterior-posterior
The combined effect of multiple mutations on protein function is hard to predict; thus, the ability to functionally assess a vast number of protein sequence variants would be practically useful for protein engineering. Here we present a high-throughput platform that enables scalable assembly and parallel characterization of barcoded protein variants with combinatorial modifications. We demonstrate this platform, which we name CombiSEAL, by systematically characterizing a library of 948 combination mutants of the widely used Streptococcus pyogenes Cas9 (SpCas9) nuclease to optimize its genome-editing activity in human cells. The ease with which the editing activities of the pool of SpCas9 variants can be assessed at multiple on- and off-target sites accelerates the identification of optimized variants and facilitates the study of mutational epistasis. We successfully identify Opti-SpCas9, which possesses enhanced editing specificity without sacrificing potency and broad targeting range. This ...
Innate Immunity has long been regarded as the non-specific arm of immune response, acting immediately and in a generic way to defend the host from infections. In the post genomic era, our knowledge of the innate immune system is enriched by findings on the specificity of innate immune reactions as well as to novel functions that do not strictly correlate with immunological defense and surveillance, immune modulation or inflammation. The advent of high-throughput platforms for genome and proteome-wide profiling, together with the enormous quantity of raw genetic information that has accumulated in the databases, have stirred new expectations in biomedical research, and led scientists to revisit established biological systems from a global and integrative perspective. Innate Immunity research now faces the challenge of integrating isolated biochemical pathways into complex gene and protein regulatory circuits. This volume collects topics on natural killer cells, mast cells, phagocytes, toll-like ...
Centrosome amplification (CA) is a hallmark of virtually all types of cancers including solid tumors and hematological malignancies. Cancer cells with extra centrosomes use centrosome clustering (CC) to allow for successful division. Because normal cells do not rely on this mechanism, CC is regarded as a promising target to selectively eradicate cells harboring supernumerary centrosomes. To identify novel inhibitors of CC, we developed a cell-based high-throughput screen that reports differential drug cytotoxicity for isogenic cell populations with different centrosome contents. We identified CP-673451 and crenolanib, two chemically related compounds originally developed for inhibition of PDGFR-β, as robust inhibitors of CC with selective cytotoxicity for cells with extra centrosomes. We demonstrate that these compounds induce mitotic spindle multipolarity by activation of the actin severing protein cofilin, leading to destabilization of the cortical actin network, and provide evidence that ...
Forward transfection is a widely used method that works well for adherent cell types; however, if one is using suspension cells or a high-throughput format, reverse transfection, where cells are added to the plated transfection reagent complex, is a more appropriate approach. This method can be used to increase the throughput and reproducibility of a CRISPR-Cas9 screen by optimizing gene editing efficiency as demonstrated in our recently published application note: "Optimization of reverse transfection of Dharmacon Edit-R synthetic crRNA and tracrRNA components with DharmaFECT transfection reagent in a Cas9-expressing cell line.". An un-cleavable ubiquitin moiety fused to EGFP allows constitutive degradation of the EGFP protein, while disruption of the proteasome components by functional protein knockout leads to accumulation of EGFP and detectable fluorescence. This functional knockout data shows that determining optimal transfection conditions prior to arrayed screening leads to high gene ...
Here we describe a rapid high-throughput method for performing RNA interference (RNAi) in moss, in which phenotyping is performed within 1 week after transformation. The moss Physcomitrella patens is a great plant model system for reverse genetic studies due to its amenability to homologous recombination as well as RNAi. Our lab has developed a rapid RNAi assay to screen for growth phenotypes in moss protonemal tissue. Here we describe how we have recently further facilitated this assay by modifying the PEG-mediated transformation protocol allowing for transformations to be carried out in a semiautomated fashion in a 96-well plate format ...
Video articles in JoVE about retinoblastoma protein include In Vivo Detection and Analysis of Rb Protein SUMOylation in Human Cells, Workflow for High-content, Individual Cell Quantification of Fluorescent Markers from Universal Microscope Data, Supported by Open Source Software, Analysis of Cell Cycle Position in Mammalian Cells, Induction of Protein Deletion Through In Utero Electroporation to Define Deficits in Neuronal Migration in Transgenic Models, Whole Mount Dissection and Immunofluorescence of the Adult Mouse Cochlea, In vitro Coculture Assay to Assess Pathogen Induced Neutrophil Trans-epithelial Migration, An In Vitro Model for Studying Cellular Transformation by Kaposi Sarcoma Herpesvirus, A Protein Preparation Method for the High-throughput Identification of Proteins Interacting with a Nuclear Cofactor Using LC-MS/MS Analysis, Genome-wide Analysis using ChIP to Identify Isoform-specific Gene Targets, Assessing Replication and Beta Cell Function in Adenovirally-transduced
Sigma-Aldrich offers abstracts and full-text articles by [Lily Mahapatra, Chengjian Mao, Neal Andruska, Chen Zhang, David J Shapiro].
The mucin MUC16 and the glycosylphosphatidylinositol anchored glycoprotein mesothelin likely facilitate the peritoneal metastasis of ovarian tumors. The biochemical basis and the kinetics of the binding between these two glycoproteins are not clearly understood. Here we have addressed this deficit and provide further evidence supporting the role of the MUC16-mesothelin interaction in facilitating cell-cell binding under conditions that mimic the peritoneal environment. In this study we utilize recombinant-Fc tagged human mesothelin to measure the binding kinetics of this glycoprotein to MUC16 expressed on the ovarian tumor cell line OVCAR-3. OVCAR-3 derived sublines that did not express MUC16 showed no affinity for mesothelin. In a flow cytometry-based assay mesothelin binds with very high affinity to the MUC16 on the OVCAR-3 cells with an apparent Kd of 5-10 nM. Maximum interaction occurs within 5 mins of incubation of the recombinant mesothelin with the OVCAR-3 cells and significant binding is