• 제목/요약/키워드: High throughput screening (HTS)

검색결과 31건 처리시간 0.024초

Establishment of High Throughput Screening System Using Human Umbilical Cord-derived Mesenchymal Stem Cells

  • Park, Eu-Gene;Cho, Tae-Jun;Oh, Keun-Hee;Kwon, Soon-Keun;Lee, Dong-Sup;Park, Seung-Bum;Cho, Jae-Jin
    • International Journal of Oral Biology
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    • 제37권2호
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    • pp.43-50
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    • 2012
  • The use of high throughput screening (HTS) in drug development is principally for the selection new drug candidates or screening of chemical toxicants. This system minimizes the experimental environment and allows for the screening of candidates at the same time. Umbilical cord-derived stem cells have some of the characteristics of fetal stem cell and have several advantages such as the ease with which they can be obtained and lack of ethical issues. To establish a HTS system, optimized conditions that mimic typical cell culture conditions in a minimal space such as 96 well plates are needed for stem cell growth. We have thus established a novel HTS system using human umbilical cord derived-mesenchymal stem cells (hUC-MSCs). To determine the optimal cell number, hUC-MSCs were serially diluted and seeded at 750, 500, 200 and 100 cells per well on 96 well plates. The maintenance efficiencies of these dilutions were compared for 3, 7, 9, and 14 days. The fetal bovine serum (FBS) concentration (20, 10, 5 and 1%) and the cell numbers (750, 500 and 200 cells/well) were compared for 3, 5 and 7 days. In addition, we evaluated the optimal conditions for cell cycle block. These four independent optimization experiments were conducted using an MTT assay. In the results, the optimal conditions for a HTS system using hUC-MSCs were determined to be 300 cell/well cultured for 8 days with 1 or 5% FBS. In addition, we demonstrated that the optimal conditions for a cell cycle block in this culture system are 48 hours in the absence of FBS. In addition, we selected four types of novel small molecule candidates using our HTS system which demonstrates the feasibility if using hUC-MSCs for this type of screen. Moreover, the four candidate compounds can be tested for stem cell research application.

정량적인 구조-활성상관 (QSAR) 기법에 의한 새로운 농약의 개발. III. 3D QSAR 기법들과 컴퓨터를 이용한 분자설계(CAMD) (Development of new agrochemicals by quantitative structure-activity relationship (QSAR) methodology. III. 3D QSAR methodologies and computer-assisted molecular design (CAMD))

  • 성낙도
    • 농약과학회지
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    • 제7권1호
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    • pp.1-11
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    • 2003
  • 새로운 농약을 탐색하고 개발하는데 있어서 고효율 유기함성 (HTOS) 기술과 고효율 검색 (HTS) 기술 등의 발전과 더불어 컴퓨터 화학을 이용한 분자설계 (CAMD) 방법으로 보편화되고 있는 비교 분자장 분석(CoMFA)과 비교 분자 유사성 지수분석(CoMSIA) 등, 3D QSAR 기법들을 위시하여 분자 홀로그램 구조 - 활성관계 (HQSAR) 분석방법 등, QSAR 기법들을 요약하고 그 활용 사례들을 간략하게 소개하였다.

Screening of Anti-Adhesion Agents for Pathogenic Escherichia coli O157:H7 by Targeting the GrlA Activator

  • Sin Young Hong;Byoung Sik Kim
    • Journal of Microbiology and Biotechnology
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    • 제33권3호
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    • pp.329-338
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    • 2023
  • Enterohemorrhagic Escherichia coli (EHEC) is a foodborne pathogen that produces attaching and effacing lesions on the large intestine and causes hemorrhagic colitis. It is primarily transmitted through the consumption of contaminated meat or fresh produce. Similar to other bacterial pathogens, antibiotic resistance is of concern for EHEC. Furthermore, since the production of Shiga toxin by this pathogen is enhanced after antibiotic treatment, alternative agents that control EHEC are necessary. This study aimed to discover alternative treatments that target virulence factors and reduce EHEC toxicity. The locus of enterocyte effacement (LEE) is essential for EHEC attachment to host cells and virulence, and most of the LEE genes are positively regulated by the transcriptional regulator, Ler. GrlA protein, a transcriptional activator of ler, is thus a potential target for virulence inhibitors of EHEC. To identify the GrlA inhibitors, an in vivo high-throughput screening (HTS) system consisting of a GrlA-expressing plasmid and a reporter plasmid was constructed. Since the reporter luminescence gene was fused to the ler promoter, the bioluminescence would decrease if inhibitors affected the GrlA. By screening 8,201 compounds from the Korea Chemical Bank, we identified a novel GrlA inhibitor named Grlactin [3-[(2,4-dichlorophenoxy)methyl]-4-(3-methylbut-2-en-1-yl)-4,5-dihydro-1,2,4-oxadiazol-5-one], which suppresses the expression of LEE genes. Grlactin significantly diminished the adhesion of EHEC strain EDL933 to human epithelial cells without inhibiting bacterial growth. These findings suggest that the developed screening system was effective at identifying GrlA inhibitors, and Grlactin has potential for use as a novel anti-adhesion agent for EHEC while reducing the incidence of resistance.

식품소재 라이브러리를 이용한 천식 완화용 물질의 초고속스크리닝 기법 개발 (Development of High Throughput Screening Techniques Using Food-borne Library against Anti-asthma Agents)

  • 허진철;박자영;권택규;정신교;김성욱;이상한
    • 한국식품저장유통학회지
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    • 제12권3호
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    • pp.267-274
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    • 2005
  • 천식과 관련하여 산화스트레스 (Oxidant stress)는 그 발병요인 중의 하나로 알려져 있다. 이에 본 연구는 농산물과 한약재를 이용하여 항산화 물질을 찾고자 하였으며, 시간과 비용의 단축을 위하여 HTS인 throughput screening)을 이용을 하였다. 항산화 실험과 관련하여서 DPPH(1,1-diphenyl-2-picrylhydrazyl), FRAP(ferric ion reducing antioxidant power), HO(hydroxyl radical) 소거, linoleic acid에 대한 항산화 활성 등을 시행하였다. 이후 $H_{2}O_2$에 의한 산화스트레스를 이용한 세포사멸을 유도하여 세포생존을 확인해 보았다. 실험결과 해바라기씨(Helianthus annuus), 신선초(Angelica utilis Makino), 시금치(Rehmannia glutinosa Libo) 등이 활성이 높게 나타났다. 본 연구는 여기에서 나온 hit를 이용하여 동물모델 실험을 진행하고자 한다.

Advances in the Development and Validation of Test Methods in the United States

  • Casey, Warren M.
    • Toxicological Research
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    • 제32권1호
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    • pp.9-14
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    • 2016
  • The National Toxicology Program Interagency Center for the Evaluation of Alternative Toxicological Methods (NICEATM) provides validation support for US Federal agencies and the US Tox21 interagency consortium, an interagency collaboration that is using high throughput screening (HTS) and other advanced approaches to better understand and predict chemical hazards to humans and the environment. The use of HTS data from assays relevant to the estrogen receptor signaling data pathway is used as an example of how HTS data can be combined with computational modeling to meet the needs of US agencies. As brief summary of US efforts in the areas of biologics testing, acute toxicity, and skin sensitization will also be provided.

Identification of a lead small-molecule inhibitor of anthrax lethal toxin by using fluorescence-based high-throughput screening

  • Wei, Dong;Bu, Zhaoyun;Yu, Ailian;Li, Feng
    • BMB Reports
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    • 제44권12호
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    • pp.811-815
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    • 2011
  • Inhalational anthrax is caused by B. anthracis, a virulent sporeforming bacterium which secretes anthrax toxins consisting of protective antigen (PA), lethal factor (LF) and edema factor (EF). LF is a Zn-dependent metalloprotease and is the main determinant in the pathogenesis of anthrax. Here we report the identification of a lead small-molecule inhibitor of anthrax lethal factor by screening an available synthetic small-molecule inhibitor library using fluorescence-based high-throughput screening (HTS) approach. Seven small molecules were found to have inhibitory effect against LF activity, among which SM157 had the highest inhibitory activity. All theses small molecule inhibitors inhibited LF in a noncompetitive inhibition mode. SM157 and SM167 are from the same family, both having an identical group complex, which is predicted to insert into S1' pocket of LF. More potent small-molecule inhibitors could be developed by modifying SM157 based on this identical group complex.

신규 제초활성 물질 발굴을 위한 메타게놈 스크리닝 방법 연구 (Establishing Effective Screening Methodology for Novel Herbicide Substances from Metagenome)

  • 이보영;최지은;김영숙;송재광;고영관;최정섭
    • Weed & Turfgrass Science
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    • 제4권2호
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    • pp.118-123
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    • 2015
  • 메타게놈(metagenome)은 연구실에서 배양이 불가능한 미생물을 포함한 자연계에 존재하는 미생물의 유전자를 직접 연구하는 학문분야이다. 지구상 거의 모든 자연, 인공 환경에서 살고 있는 미생물 DNA를 분리 정제하는 것이 가능하며, 재조합 DNA 기술 등을 이용하여 배양 가능한 숙주미생물에 메타지놈을 클로닝함으로서 메타지놈 라이브러리를 제작할 수 있다. 최근 메타게노믹스를 통하여 자연계에 존재하고 있는 대다수의 미생물들이 실험실에서 배양되지 않았던 이유를 구명할 수 있게 되었고, 그들이 가지고 있는 생태학적인 의미와 역할에 대한 이해와 더불어 점차 그 응용 분야와 범위도 확대되고 있다. 이와 같은 메타게놈의 응용 분야 확대의 한 방안으로 본 연구에서는 새로운 제초활성 물질 및 제초활성 물질 생산에 필요한 유전자를 확보하기 위해 메타게놈 라이브러리를 대상으로 오이 떡잎절편 검정, 미세조류 생장저해 검정, 종자발아 저해 검정의 HTS (high throughput screening) 시스템을 구축하였고, 구축된 시스템으로부터 선발된 최종 단일 클론인 9-G1과 9-G12의 바랭이에 대한 in vivo assay를 통해 본 연구에서 개발한 HTS 시스템의 유효성을 확인 하였다. 후속연구로서 활성단일클론이 만들어내는 제초활성물질의 동정 및 제초활성물질을 합성하는 유전자군에 대한 연구를 수행 중에 있다.

Design and Combonatorial Synthesis of Selenoflavonoids and Indole Compounds for Neuroprotective Drug

  • Jeong, Jin-Hyun
    • 대한약학회:학술대회논문집
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    • 대한약학회 2003년도 Proceedings of the Convention of the Pharmaceutical Society of Korea Vol.1
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    • pp.69-71
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    • 2003
  • As recent as 10 years ago, a method of developing new medicine was developed by a new compounding method moving away from an existing one. Combinatorial chemistry made it easier to combine various kinds of compounds in a very short time and with little effort from existing methods. Through combinatorial chemistry, a number of compounds were synthesized using HTS(High Throughput Screening), with many reports reaching a clinical stage in search of new candidate material. (omitted)

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Systemic and Cell-Type Specific Profiling of Molecular Changes in Parkinson's Disease

  • Lee, Yunjong
    • Interdisciplinary Bio Central
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    • 제4권3호
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    • pp.6.1-6.12
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    • 2012
  • Parkinson's disease (PD) is a complicated neurodegenerative disorder although it is oftentimes defined by clinical motor symptoms originated from age dependent and progressive loss of dopaminergic neurons in the midbrain. The pathogenesis of PD involves dopaminergic and nondopaminergic neurons in many brain regions and the molecular mechanisms underlying the death of different cell types still remain to be elucidated. There are indications that PD causing disease processes occur in a global scale ranging from DNA to RNA, and proteins. Several PD-associated genes have been reported to play diverse roles in controlling cellular functions in different levels, such as chromatin structure, transcription, processing of mRNA, translational modulation, and posttranslational modification of proteins. The advent of quantitative high throughput screening (HTS) tools makes it possible to monitor systemic changes in DNA, RNA and proteins in PD models. Combined with dopamine neuron isolation or derivation of dopamine neurons from PD patient specific induced pluripotent stem cells (PD iPSCs), HTS techonologies will provide opportunities to draw PD causing sequences of molecular events in pathologically relevant PD samples. Here I discuss previous studies that identified molecular functions in which PD genes are involved, especially those signaling pathways that can be efficiently studied using HTS methodologies. Brief descriptions of quantitative and systemic tools looking at DNA, RNA and proteins will be followed. Finally, I will emphasize the use and potential benefits of PD iPSCs-derived dopaminergic neurons to screen signaling pathways that are initiated by PD linked gene mutations and thus causative for dopaminergic neurodegneration in PD.

High Throughput 프로세스에서 품질혁신의 성능평가를 위한 Z-Factor의 적용방안 (Implementation of Z-Factor Statistics for Performance Evaluation of Quality Innovation in the High Throughput Process)

  • 최성운
    • 대한안전경영과학회지
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    • 제15권1호
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    • pp.293-301
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    • 2013
  • The purpose of this study is to introduce the limit of previously used six sigma quality process evaluation metrics, $Z_{st}$ and $P_{pk}$, and a solution to overcome this drawback by using a metric based on performance evaluation of Z-factor quality innovation. Case analysis on projects from national six sigma contest from 2011 to 2012 is performed and literature review on new drug development HTS (High Throughput Screening) is used to propose innovative performance evaluation metrics. This research shows that experimental study on six sigma evaluation metric, $Z_{st}$ and $P_{pk}$, have no significance difference between industrial type (Manufacturing, Semi-Public Institute, Public Institute) and CTQ type (Product Technology Type CTQ, Process Technology Type CTQ). Following discovery characterize this quality improvement as fixed target type project. As newly developed moving target type of quality innovation performance metric Z-Factor is used for evaluating experimental study, hypothetical analysis suggests that $Z_{st}$ and $P_{pk}$ share different relationship or even show reciprocal relationship. Constraints of the study are relatively small sample size of only 37 projects from past 2 years and conflict on having interview and communication with six sigma quality practitioner for qualitative experimental study. Both moving target type six sigma innovation project and fixed target type improvement project or quality circle enables efficient ways for a better understanding and quality practitioner use by applying quality innovation performance metric. Downside of fixed target type quality performance evaluation metric, $Z_{st}$ and $P_{pk}$, is presented through experimental study. In contrast, advantage of this study is that high throughput requiring product technology, process technology and quantum leap typed innovation effect is evaluated based on precision and accuracy and Z-Factor that enables relative comparison between enterprises is proposed and implemented.