1,721,029 research outputs found
Development of high content image-based cell viability assays
Full text linkAim and applications: 1. Optimize fluorescence-based cellular assays for getting insights about mechanisms leading to cell death 2. Informative high content assays development with the aim to implement them in order to obtain preliminary toxicity information of compounds 3. Toxicity profiling of chemicals or biologicals used in screening, in particular hits behavior relative to the characterization of cell viability. Methods: Different fluorescent probes for cell viability were tested and optimized in HeLa cells by varying parameters of staining, controls and incubation times. Automated fluorescence microscopy was used to image cells in 96-well plates and automated image analysis provided tool for quantification of the response to the different assays. Image quality was estimated in each condition and ability to discriminate between positive and negative populations was statistically determined after image segmentation and features extraction. Results: Live/dead assay using calcein-AM and ethidium homodimer-1 showed significant difference in signal between living and dead cells but high variability was observed within the same population. Apoptosis probes FLICA and annexin V conjugate were tested on cells treated with staurosporine. FLICA staining gave low signal-to-background ratio and apoptotic population was not significantly discernible. Living, early apoptotic and necrotic/late apoptotic cells populations were segregated using annexin V assay. Autophagy detection using LC3B immunostaining was observable in cells treated with chloroquine. Developed image analysis method reliably segmented autophagosomes in autophagic cells but failed with dead cells. Assays were tested on a set of toxic compounds and results showed differences with information found on these substances. Conclusion: Fluorescent probes are a powerful tool to assess cell death mechanism in high content screening assay as they can report multiple biological activities at the same time. As information in this type of assay are gathered at a single cell level, high variability is expected and must be reduced as possible to successfully characterize the phenotypes encountered and reach screening requirements.SSVBiomolecular Screening Facility, EPF
Cellular transfection of small interfering ribonucleic acids. Electroporation as an alternative to liposome-based delivery transfection methods
No full textRNA interference has become an increasingly important tool for all aspects of molecular biology. Nevertheless, there is a technological challenge for performing gene knock-down with chemical transfection agents, as many relevant cell types are refractory to efficient transport of short interfering RNA (siRNA) into cytosol using these classical carrier-based methods. The aim of this project was to evaluate the potential of electroporation as a generic transfection method and to compare it with a standard and well-established chemical reversetransfection using cationic lipids as carriers for short-interfering RNAs. We have used an automated cell electroporation instrument from a swedish company, Cellectricon. The Cellaxess HT instrument is able to perform electroporation in 384 wells plates. We compared cell viability, transfection efficiency and extend of gene silencing caused by electroporation, with reverse-transfection using Lipofectamine RNAiMAX. Gene knock down efficiency has been measured at the protein function level by targeting ubiquitously expressed genes that are essential to cell survival. Cell viability and transfection efficiency are, then, opposed with this strategy. Results clearly showed a net improvement of "difficult-to-transfect" cell transfection but this improvement comes at the price of consuming up to ten fold more siRNAs. For "normal" cell line, lipofection remains the preferred methodology as it is more efficient and less expensive. Furthermore, the Cellaxess HT instrument has some serious drawbacks that pervert easy and reproducible result acquisition.SSVBiomolecular Screening Facility, EPF
Multiplexed detection of nucleic acids - testing and benchmarking of a novel platform developed by Biocartis
No full textSSVBiomolecular Screening Facility EPFL. - Carried out in the company Biocartis located at PSE of the EPFL, under the supervision of Elisa Leimgruber & Didier Falconne
Going Beyond Counting First Authors in Author Co-citation Analysis
Full text linkThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
Variations on the Author
Full text link“Variations on the Author” discusses two of Eduardo Coutinho’s recent films (Um Dia na Vida, from 2010, and Últimas Conversas, posthumously released in 2015) and their contribution to the general question of documentary authorship. The director’s filmography is characterized by a consistent yet self-effacing form of authorial self-inscription: Coutinho often features as an interviewer that rather than express opinions propels discourses; an interviewer that is good at listening. This mode of self-inscription characterizes him as an author who is not expressive but who is nonetheless markedly present on the screen. In Um Dia na Vida, however, Coutinho is completely absent form the image, while Últimas Conversas, on the contrary, includes a confessional prologue that moves the director from the margins to the center of his films. This article examines the ways in which these works stand out in the filmography of a director who offers new insights into the notion of cinematic authorship
Novel fluorescence - based approaches to probe ligand recognition and structure of the tachykinin NK2 receptor
No full textLCPP
Appropriate Similarity Measures for Author Cocitation Analysis
Full text linkWe provide a number of new insights into the methodological discussion about author cocitation analysis. We first argue that the use of the Pearson correlation for measuring the similarity between authors’ cocitation profiles is not very satisfactory. We then discuss what kind of similarity measures may be used as an alternative to the Pearson correlation. We consider three similarity measures in particular. One is the well-known cosine. The other two similarity measures have not been used before in the bibliometric literature. Finally, we show by means of an example that our findings have a high practical relevance.information science;Pearson correlation;cosine;similarity measure;author cocitation analysis
Developing the Biomolecular Screening Facility at the EPFL into the Chemical Biology Screening Platform for Switzerland
Full text linkThe Biomolecular Screening Facility (BSF) is a multidisciplinary laboratory created in 2006 at the Ecole Polytechnique Federale de Lausanne (EPFL) to perform medium and high throughput screening in life sciences-related projects. The BSF was conceived and developed to meet the needs of a wide range of researchers, without privileging a particular biological discipline or therapeutic area. The facility has the necessary infrastructure, multidisciplinary expertise and flexibility to perform large screening programs using small interfering RNAs (siRNAs) and chemical collections in the areas of chemical biology, systems biology and drug discovery. In the framework of the National Centres of Competence in Research (NCCR) Chemical Biology, the BSF is hosting 'ACCESS', the Academic Chemical Screening Platform of Switzerland that provides the scientific community with chemical diversity, screening facilities and know-how in chemical genetics. In addition, the BSF started its own applied research axes that are driven by innovation in thematic areas related to preclinical drug discovery and discovery of bioactive probes.PTC
Quantification of stored red blood cell fluctuations by time-lapse holographic cell imaging
Full text linkWe propose methods to quantitatively calculate the fluctuation rate of red blood cells with nanometric axial and millisecond temporal sensitivity at the single-cell level by using time-lapse holographic cell imaging. For this quantitative analysis, cell membrane fluctuations (CMFs) were measured for RBCs stored at different storage times. Measurements were taken over the whole membrane for both the ring and dimple sections separately. The measurements show that healthy RBCs that maintain their discocyte shape become stiffer with storage time. The correlation analysis demonstrates a significant negative correlation between CMFs and the sphericity coefficient, which characterizes the morphological type of erythrocyte. In addition, we show the correlation results between CMFs and other morphological properties such as projected surface area, surface area, mean corpuscular volume, and mean corpuscular hemoglobin. (C) 2018 Optical Society of America under the terms of the OSA Open Access Publishing AgreementPTC
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