Journal of Biomolecular Screening最新文献_第2页

2009 Meetings Calendar 2009年会议日程表

Journal of Biomolecular Screening Pub Date : 2009-01-01 DOI: 10.1177/1087057108330291

引用次数: 0

A fluorescence lifetime-based assay for protease inhibitor profiling on human kallikrein 7. 基于寿命的蛋白酶抑制剂谱分析的荧光分析。

Journal of Biomolecular Screening Pub Date : 2009-01-01 DOI: 10.1177/1087057108327328

Klaus Doering, Gabriele Meder, Manuela Hinnenberger, Julian Woelcke, Lorenz M Mayr, Ulrich Hassiepen

{"title":"A fluorescence lifetime-based assay for protease inhibitor profiling on human kallikrein 7.","authors":"Klaus Doering, Gabriele Meder, Manuela Hinnenberger, Julian Woelcke, Lorenz M Mayr, Ulrich Hassiepen","doi":"10.1177/1087057108327328","DOIUrl":"https://doi.org/10.1177/1087057108327328","url":null,"abstract":"Fluorescence lifetime is an intrinsic parameter describing the fluorescence process. Changes in the fluorophore's physicochemical environment can lead to changes in the fluorescence lifetime. When used as the readout in biological assays, it is thought to deliver superior results to conventional optical readouts. Hence it has the potential to replace readout technologies currently established in drug discovery such as absorption, luminescence or fluorescence intensity. Here we report the development of an activity assay for human kallikrein 7, a serine protease involved in skin diseases. As a probe, we have selected a blue-fluorescent acridone dye, featuring a remarkably long lifetime that can be quenched by either of the 2 natural amino acids, tyrosine and tryptophan. Incorporating this probe and 1 of the quenching amino acids on either side of the scissile bond of the substrate peptide enables us to monitor the enzymatic activity by quantifying the increase in the fluorescence lifetime signal. A systematic investigation of substrate structures has led to a homogenous, microplate-based, compound profiling assay that yields inhibitory constants down into the single-digit nanomolar range. This type of assay has now been added to our standard portfolio of screening techniques, and is routinely used for compound profiling.","PeriodicalId":15087,"journal":{"name":"Journal of Biomolecular Screening","volume":"14 1","pages":"1-9"},"PeriodicalIF":0.0,"publicationDate":"2009-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1177/1087057108327328","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"27948090","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 26

Development and validation of a higher throughput screening approach to genotoxicity testing using the GADD45a-GFP GreenScreen HC assay. 利用 GADD45a-GFP GreenScreen HC 检测法，开发并验证了基因毒性检测的高通量筛选方法。

Journal of Biomolecular Screening Pub Date : 2009-01-01 DOI: 10.1177/1087057108327065

Andrew W Knight, Louise Birrell, Richard M Walmsley

{"title":"Development and validation of a higher throughput screening approach to genotoxicity testing using the GADD45a-GFP GreenScreen HC assay.","authors":"Andrew W Knight, Louise Birrell, Richard M Walmsley","doi":"10.1177/1087057108327065","DOIUrl":"10.1177/1087057108327065","url":null,"abstract":"There is a pressing need to develop rapid yet accurate screening assays for the identification of genotoxic liability and for early hazard assessment in drug discovery. The GADD45a-GFP human cell-based genotoxicity assay (GreenScreen HC) has been reformatted to test 12 compounds per 96-well microplate in a higher throughput, automated screening mode and the protocol applied to the analysis of 1266 diverse, pharmacologically active compounds. Testing from a fixed starting concentration of 100 AmicroM and over 3 serial dilutions, the hit rates for genotoxicity (7.3%) and cytotoxicity (33%) endpoints of the assay have been determined in a much wider chemical space than previously reported. The degree of interference from color, autofluorescence, and low solubility has also been assessed. The assay results have been compared to an in silico approach to genotoxicity assessment using Derek for Windows software. Where carcinogenicity data were available, GreenScreen HC demonstrated a higher specificity than in silico methods while identifying genotoxic species that were not highlighted for genotoxic liability in structure-activity relationship software. Higher throughput screening from a fixed, low concentration reduces sensitivity to less potent genotoxins, but the maintenance of the previously reported high specificity is essential in early hazard assessment where misclassification can lead to the needless rejection of potentially useful compounds in drug development.","PeriodicalId":15087,"journal":{"name":"Journal of Biomolecular Screening","volume":"14 1","pages":"16-30"},"PeriodicalIF":0.0,"publicationDate":"2009-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1177/1087057108327065","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"27948092","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 27

Process validation and screen reproducibility in high-throughput screening. 高通量筛选中的工艺验证和筛选再现性。

Journal of Biomolecular Screening Pub Date : 2009-01-01 DOI: 10.1177/1087057108326664

Isabel Coma, Liz Clark, Emilio Diez, Gavin Harper, Jesus Herranz, Glenn Hofmann, Mark Lennon, Nicola Richmond, Manuel Valmaseda, Ricardo Macarron

{"title":"Process validation and screen reproducibility in high-throughput screening.","authors":"Isabel Coma, Liz Clark, Emilio Diez, Gavin Harper, Jesus Herranz, Glenn Hofmann, Mark Lennon, Nicola Richmond, Manuel Valmaseda, Ricardo Macarron","doi":"10.1177/1087057108326664","DOIUrl":"https://doi.org/10.1177/1087057108326664","url":null,"abstract":"The use of large-scale compound screening has become a key component of drug discovery projects in both the pharmaceutical and the biotechnological industries. More recently, these activities have also been embraced by the academic community as a major tool for chemical genomic activities. High-throughput screening (HTS) activities constitute a major step in the initial drug discovery efforts and involve the use of large quantities of biological reagents, hundreds of thousands to millions of compounds, and the utilization of expensive equipment. All these factors make it very important to evaluate in advance of the HTS campaign any potential issues related to reproducibility of the experimentation and the quality of the results obtained at the end of these very costly activities. In this article, the authors describe how GlaxoSmithKline (GSK) has addressed the need of a true validation of the HTS process before embarking in full HTS campaigns. They present 2 different aspects of the so-called validation process: (1) optimization of the HTS workflow and its validation as a quality process and (2) the statistical evaluation of the HTS, focusing on the reproducibility of results and the ability to distinguish active from nonactive compounds in a vast collection of samples. The authors describe a variety of reproducibility indexes that are either innovative or have been adapted from generic medical diagnostic screening strategies. In addition, they exemplify how these validation tools have been implemented in a number of case studies at GSK.","PeriodicalId":15087,"journal":{"name":"Journal of Biomolecular Screening","volume":"14 1","pages":"66-76"},"PeriodicalIF":0.0,"publicationDate":"2009-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1177/1087057108326664","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"27948552","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 42

Comparison of variability and sensitivity between nuclear translocation and luciferase reporter gene assays. 核易位和荧光素酶报告基因检测的变异性和敏感性比较。

Journal of Biomolecular Screening Pub Date : 2009-01-01 DOI: 10.1177/1087057108328016

Vincent Unterreiner, Yvonne Ibig-Rehm, Marjo Simonen, Hanspeter Gubler, Daniela Gabriel

{"title":"Comparison of variability and sensitivity between nuclear translocation and luciferase reporter gene assays.","authors":"Vincent Unterreiner, Yvonne Ibig-Rehm, Marjo Simonen, Hanspeter Gubler, Daniela Gabriel","doi":"10.1177/1087057108328016","DOIUrl":"https://doi.org/10.1177/1087057108328016","url":null,"abstract":"High-content screening (HCS), a technology based on subcellular imaging by automated microscopy and sophisticated image analysis, has emerged as an important platform in small-molecule screening for early drug discovery. To validate a subcellular imaging assay for primary screening campaigns, an HCS assay was compared with a non-image-based readout in terms of variability and sensitivity. A study was performed monitoring the accumulation of the forkhead transcription factor of the O subfamily (FOXO3a) coupled with green fluorescent protein in the nucleus of human osteosarcoma (U-2 OS) cells. In addition, the transcription of a luciferase gene coupled with a FOXO3a-responsive promoter was monitored. This report demonstrates that both assay formats show good reproducibility in primary and concentration response screening despite differences in statistical assay quality. In primary screening, the correlation of compound activity between the 2 assays was low, in contrast to the good correlation of the IC(50) values of confirmed compounds. Furthermore, the high-content imaging assay showed a mean shift of 2.63-fold in IC(50) values compared with the reporter gene assay. No chemical scaffold was specifically found with 1 of the technologies only, however these results validate the HCS technology against established assays for screening of new molecular entities.","PeriodicalId":15087,"journal":{"name":"Journal of Biomolecular Screening","volume":"14 1","pages":"59-65"},"PeriodicalIF":0.0,"publicationDate":"2009-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1177/1087057108328016","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"27948551","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 8

Mouse Bsep ATPase assay: a nonradioactive tool for assessment of the cholestatic potential of drugs. 小鼠Bsep三磷酸腺苷酶测定:一种评估药物抑胆潜能的非放射性工具。

Journal of Biomolecular Screening Pub Date : 2009-01-01 Epub Date: 2008-11-21 DOI: 10.1177/1087057108326145

Emese Kis, Zsuzsanna Rajnai, Eniko Ioja, Krisztina Herédi Szabó, Tünde Nagy, Dóra Méhn, Péter Krajcsi

{"title":"Mouse Bsep ATPase assay: a nonradioactive tool for assessment of the cholestatic potential of drugs.","authors":"Emese Kis, Zsuzsanna Rajnai, Eniko Ioja, Krisztina Herédi Szabó, Tünde Nagy, Dóra Méhn, Péter Krajcsi","doi":"10.1177/1087057108326145","DOIUrl":"https://doi.org/10.1177/1087057108326145","url":null,"abstract":"The mouse ortholog of the human bile salt export pump (BSEP) transporter was expressed in a baculovirus-infected insect cell (Sf9) system to study the effect of membrane cholesterol content on the transporter function. The transport activity of cholesterol-loaded mouse Bsep-HAM-Sf9 vesicles was determined in a vesicular transport assay with taurochenodeoxycholate (TCDC), a known BSEP substrate. Mouse Bsep transports TCDC at a high rate that can be sensitively detected in the ATPase assay. Cholesterol upload of the Sf9 membrane potentiates both TCDC transport and TCDC-stimulated ATPase activities. Inhibitory effect of BSEP interactors on probe substrate transport was tested in both vesicular transport and ATPase assays using cholesterol-loaded membrane vesicles. A good rank order correlation was found between IC(50) values measured in TCDC-stimulated mBsep ATPase assay and in the human BSEP vesicular transport assay utilizing taurocholate (TC) as probe substrate. This upgraded form of the mouse Bsep-HAM ATPase assay is a user friendly, sensitive, nonradioactive method for early high-throughput screening of drugs with BSEP-related cholestatic potential. It may complement the human BSEP-mediated taurocholate vesicular transport inhibition assay.","PeriodicalId":15087,"journal":{"name":"Journal of Biomolecular Screening","volume":"14 1","pages":"10-5"},"PeriodicalIF":0.0,"publicationDate":"2009-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1177/1087057108326145","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"27855422","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 19

Product Focus 产品集中

Journal of Biomolecular Screening Pub Date : 2008-10-01 DOI: 10.1177/10870571080130091501

引用次数: 0

Product Focus 产品集中

Journal of Biomolecular Screening Pub Date : 2008-09-01 DOI: 10.1177/10870571080130081201

引用次数: 0

Product Focus: Screening Robotics and Automation 产品重点:筛选机器人及自动化

Journal of Biomolecular Screening Pub Date : 2008-07-01 DOI: 10.1177/10870571080130061201

引用次数: 0

Book Review: Immunoassay and Other Bioanalytical Techniques, edited by Jeanette M. Van Emon. Boca Raton, FL: CRC Press, Taylor & Francis Group; 2007. 512 pp. Hardcover. ISBN 10:0—8493—3942—1 书评:免疫测定和其他生物分析技术，由Jeanette M. Van Emon编辑。博卡拉顿，佛罗里达州:CRC出版社，泰勒和弗朗西斯集团;2007. 512页，精装。ISBN 10:0 - 8493 - 3942 - 1

Journal of Biomolecular Screening Pub Date : 2008-06-01 DOI: 10.1177/1087057108317441

T. Chung

引用次数: 1