Flow Cytometry CORE

流式细胞术核心

• 批准号: 10266926
• 负责人: Rachel R. Caspi
• 金额: $ 56.45万
• 依托单位: NATIONAL EYE INSTITUTE
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10266926
• 关键词: 2 year old; Age; Aging; Apoptosis; Blood; Budgets; COVID-19 pandemic; Cells; Collaborations; Communities; Computer software; Cytometry; DNA; DNA Methylation; Data Analyses; Detection; Education; Energy Metabolism; Equipment; Eye; Flow Cytometry; Foundations; Gene Expression; Genetic Transcription; Genotype; Grant; Hour; Human; Information Technology; Interleukins; Intramural N.I.H. Research Support; Investments; Laboratories; Leukocytes; Liquid substance; Measurement; Membrane; Methods; Monitor; National Eye Institute; National Institute of Allergy and Infectious Disease; Peripheral Blood Mononuclear Cell; Phenotype; Ploidies; Postdoctoral Fellow; Preparation; Principal Investigator; Procedures; Production; Proteins; Protocols documentation; Publications; Publishing; Reporter Genes; Research; Research Personnel; Research Project Grants; Resources; Sampling; Science; Secure; Services; Sorting - Cell Movement; Source; Specimen; Stains; Students; Supervision; Techniques; Tissues; Training; United States National Institutes of Health; Work; autoimmune uveitis; coronavirus disease; cytokine; data acquisition; equipment acquisition; exosome; genome-wide; human tissue; instrument; instrumentation; neuroinflammation; operation; recruit; retinal rods; square foot; volunteer

GENERAL OVERVIEW: The Flow Cytometry Core at NEI provides flow cytometry analytical and sorting equipment and services to the NEI Intramural community. It utilizes and develops state-of-the-art sample preparation, data acquisition and analysis, and sorting procedures in collaborative research projects. Provides training to students, fellows, and principal investigators on sample preparation, staining, and post-sort handling. Assesses technical research needs and recommends recruitment of the appropriate staff and acquisition of the equipment needed to meet those needs. The core also supports NIH intramural research outside NEI by processing samples for laboratories without access to flow cytometry instrumentation and collaborations with the Foundation for Advanced Education in the Sciences. The work of the cytometry core was tremendously impacted by the emergence of the COVID-19 pandemic. The laboratory was secured and closed in March 2020. In May 2020, the core partially restarted for a Covid related project. In August, the biologist in the core moved to NIAID. As of September, users of the core have not been able to restart their regular operations. SERVICES PROVIDED THE CORE: This year, fifty-fivindividuals from twentyfour different laboratories used the facility. These services and collaborative services were performed for 10 Principal Investigators (PIs) from 5 NEI labs (ERPD, LI, LRCMB, MSF, N-NRL, RN, OGVFB and UNGIRD), plus 15 other laboratories at NIH. This year the core performed 750 hours of sorting and over 9,000 samples were analyzed. Among the techniques now in use within the core are methods for phenotyping live cells, gene expression detection through fluorescent reporter genes, monitoring membrane and DNA content changes due to apoptosis or proliferation, measurement of intracellular proteins and quantification of soluble proteins. The work involving human tissues includes the sorting of peripheral blood mononuclear cells to study their cytokine production, genotype and DNA or RNA expression. The sources are blood, buffy coat, and white cells. Some analytical work had been done with eye fluids, eye tissue specimen, protein, and tears. The National Eye Institute made a great investment in biosafety with the addition BD FACSAria Fusion flow cytometer equipped with a fully integrated biosafety cabinet. This sorter meets the recent NIH's operator and sample protection requirements as well as global standards for bioprotection for processing human samples. No human tissues were stored by the core. The Core encourages, but does not require, users to acknowledge the Core contribution in their publications. The following are examples of the publications that used NEI Flow Cytometry Core resources: Corso-Daz X, Gentry J, Rebernick R, et al. Genome-wide Profiling Identifies DNA Methylation Signatures of Aging in Rod Photoreceptors Associated with Alterations in Energy Metabolism. Cell Rep. 2020;31(3):107525. doi:10.1016/j.celrep.2020.107525 Kang M, Choi JK, Jittayasothorn Y, Egwuagu CE. Interleukin 35-Producing Exosomes Suppress Neuroinflammation and Autoimmune Uveitis. Front Immunol. 2020;11:1051. Published 2020 May 29. doi:10.3389/fimmu.2020.01051 TRAINING: Several formal training sessions were offered by the Core to the NEI community. These courses included: Introduction to Flow Cytometry, Advanced Techniques (Ex. Apoptosis applications), Analytical Instrument Operation and Data Analysis with FloJo Software. The Core stuff received about 100 hrs. or training a year in flow cytometry. The Core stuff provided about 50 hrs of training to the NEI community and 60 hrs of training through FAES to the NIH community.

总体概述： NEI 的流式细胞术核心为 NEI 校内社区提供流式细胞术分析和分选设备及服务。它在合作研究项目中利用和开发最先进的样品制备、数据采集和分析以及分类程序。为学生、研究员和主要研究人员提供样品制备、染色和分选后处理方面的培训。评估技术研究需求并建议招聘适当的人员并采购满足这些需求所需的设备。该核心还支持 NEI 之外的 NIH 校内研究，为无法使用流式细胞仪的实验室处理样本，并与科学高级教育基金会合作。 COVID-19 大流行的出现对细胞计数核心的工作产生了巨大影响。该实验室于 2020 年 3 月安全并关闭。2020 年 5 月，核心部分重启 Covid相关项目。 8月，核心生物学家转到NIAID。截至 9 月份，核心用户还无法重新启动其常规操作。 提供的服务核心： 今年，来自二十四个不同实验室的五十五个人使用了该设施。这些服务和协作服务是为来自 5 个 NEI 实验室（ERPD、LI、LRCMB、MSF、N-NRL、RN、OGVFB 和 UNGIRD）以及 NIH 的其他 15 个实验室的 10 名首席研究员 (PI) 提供的。今年，核心进行了 750 个小时的分选，分析了 9,000 多个样本。 目前核心使用的技术包括活细胞表型分析方法、通过荧光报告基因检测基因表达、监测细胞凋亡或增殖导致的膜和 DNA 含量变化、测量细胞内蛋白质以及定量可溶性蛋白质。 涉及人体组织的工作包括分选外周血单核细胞，以研究其细胞因子的产生、基因型以及 DNA 或 RNA 表达。来源是血液、血沉棕黄层和白细胞。对眼液、眼组织样本、蛋白质和眼泪进行了一些分析工作。美国国家眼科研究所在生物安全方面进行了大量投资，新增了配备完全集成生物安全柜的 BD FACSAria Fusion 流式细胞仪。该分选机满足最近 NIH 的操作员和样品保护要求以及处理人体样品的生物保护全球标准。核心中没有储存任何人体组织。 核心鼓励但不要求用户在其出版物中承认核心的贡献。以下是使用 NEI 流式细胞术核心资源的出版物示例： Corso-Daz X、Gentry J、Rebernick R 等人。全基因组分析鉴定了与能量代谢改变相关的视杆光感受器衰老的 DNA 甲基化特征。细胞报告。2020；31(3):107525。 doi:10.1016/j.celrep.2020.107525 Kang M、Choi JK、Jittayasothorn Y、Egwuagu CE。产生白细胞介素 35 的外泌体可抑制神经炎症和自身免疫性葡萄膜炎。前免疫学。 2020；11：1051。发布于 2020 年 5 月 29 日。doi:10.3389/fimmu.2020.01051 训练： 核心向 NEI 社区提供了几次正式培训课程。这些课程包括：流式细胞术简介、先进技术（例如细胞凋亡应用）、分析仪器操作和使用 FloJo 软件进行数据分析。 核心内容花了大约 100 小时。或接受一年的流式细胞术培训。 核心人员为 NEI 社区提供了大约 50 小时的培训，并通过 FAES 为 NIH 社区提供了 60 小时的培训。