ShEEP Request for Cellular & Molecular Analysis Core

ShEEP 移动电话请求

• 批准号: 9905897
• 负责人: Jeffrey A. Jones
• 金额: --
• 依托单位: RALPH H JOHNSON VA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-01 至 2020-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9905897
• 关键词: Address; Affect; Agreement; Animal Model; Animals; Area; Biochemical; Biological; Blood; Blood Cells; Caring; Cells; Complement; Copy Number Polymorphism; Core Facility; DNA; Data Quality; Detection; Diagnosis; Disease; Drops; Equipment; Evaluation; Evidence based treatment; Fluorescent Antibody Technique; Functional disorder; Funding; Gel; Gene Expression Profiling; Genetic Diseases; Genotype; Healthcare; Heart Diseases; Human; Image Analysis; Individual; Infrastructure; Injury; Investigation; Laboratories; Maintenance; Medical; Medical center; Mental disorders; Methods; MicroRNAs; Military Personnel; Mission; Modeling; Molecular; Molecular Analysis; Musculoskeletal; Nature; Neurologic; Orthopedics; Pathology; Patients; Proteins; Protocols documentation; RNA; Reproducibility; Research; Research Personnel; Rodent; Sampling; Services; Sheep; Specimen; Spinal cord injury; Stains; Stroke; Techniques; Technology; Time; Tissues; Transcript; Translating; Traumatic Brain Injury; Untranslated RNA; Validation; Vascular Diseases; Veterans; Western Blotting; Work; X-Ray Film; circular RNA; circulating biomarkers; clinical practice; cost; design; digital; experience; fluorescence imaging; human disease; imaging system; instrumentation; next generation sequencing; operation; outcome forecast; programs; tool

This proposal addresses a need for the enhancement of our Cell and Molecular Evaluation Core (CMEC) with the addition of digital drop PCR (ddPCR) and chemiluminescence/fluorescence imaging analysis; two technologies that will enhance our ability to complete every day, as well as, advanced analyses of blood, cells, and tissues, as a part of the common work flow associated with our individually funded projects. Bio-Rad's ddPCR and highly sensitive ChemiDoc MP imaging systems, provide state-of-the-art analyses that will accelerate research efforts in areas important to the VA medical mission including mental health disorders, traumatic brain injury, spinal cord injury, cardiac disease, orthopedic injury, and stroke. Furthermore, these cell and molecular analysis tools will complement existing technology in the Cell and Molecular Evaluation Core housed within the Research Service at the Ralph H. Johnson VA Medical Center in Charleston, SC. We aim to provide state-of-the-art equipment to our investigators, in order to facilitate comprehensive methods of evaluation spanning from the subcellular level to the whole animal. These core facilities, built around key equipment items often costing more than what a single laboratory can provide, have been established by design in order to support and advance our wide-ranging research program that directly impacts the health care of our Veterans. The central hypothesis addressed by this core, is that advanced technology that provides a highly sensitive assessment of disease or injury presence and progression at the molecular level, will enhance the biological significance of the work being done by our VA investigators. This hypothesis will be addressed through 4 Primary Analytical Objectives: 1) Highly sensitive detection and absolute quantitation of DNA and RNA targets as low as 1 out of 1,000,000 total copies. Bio-Rad's ddPCR is highly suited for the study of low copy number transcripts from biological specimens that are limited in size (rodent tissues, etc.), or exist at very low abundance, and represent targets that are often critical for the detection and analysis of human disease (i.e. the examination of circulating biomarkers like micro-RNAs, circular-RNAs, long non-coding-RNAs) and that can be used for diagnosis and prognosis; 2) Standard DNA and RNA analytical techniques commonly used in a versatile and scalable workflow (i.e. genotyping, gene expression analysis, copy number variation, and Next-Generation- Sequencing validation); 3) Reproducible quantitation and normalization of protein bands on Western blots (to total protein per lane with Bio-Rad stain-free gel technology) using standard chemiluminescence protocols and a highly sensitive imaging system, in place of x-ray film; and 4) Multiplex fluorescent antibodies detection, in an effort to identify multiple analytes simultaneously by western blotting. This not only cuts down on time-to-results, but also on usage of consumables. Given the versatility and integrated nature of our core facilities, VA Investigators will be able obtain high-quality data, from “every-day” and advanced technical protocols, in a scalable fashion, in order to facilitate the biochemical analysis of blood, cells, and tissues as a critical component of their funded studies. The results obtained from the examination of DNA, RNA, and protein, are directly applicable to the pathophysiology of the core medical issues affecting our Veterans such as vascular disease, cardiac disease, neurologic illness, musculoskeletal pathologies, stroke, traumatic brain injury, and spinal cord injury. The integration of the proposed equipment with our existing Cell and Molecular Evaluation Core will fill technical gaps and allow our researchers to conduct full-service biochemical assessments of their samples. This represents a unique infrastructure that will significantly advance the research being conducted at our local VAMC. The requested instrumentation will further enhance local VA research capabilities in line with the objectives outlined within the Secretary of the Department of Veterans Affairs Blueprint for Excellence.

该提案解决了对我们的细胞增强和分子评估核心（CMEC）的需求 添加数字滴PCR（DDPCR）和化学发光/荧光成像分析；二 将增强我们每天完成的能力以及对血液，细胞，高级分析的技术 和组织，作为与我们个人资助的项目相关的共同工作流程的一部分。 Bio-Rad的 DDPCR和高度敏感的Chemidoc MP成像系统，提供最先进的分析 加快对VA医疗任务至关重要的领域的研究工作，包括心理健康障碍， 脑外伤，脊髓损伤，心脏病，骨科损伤和中风。此外，这些细胞 分子分析工具将在细胞和分子评估核心中完成现有技术 位于南卡罗来纳州查尔斯顿的Ralph H. Johnson VA医疗中心的研究服务中。我们的目标 为我们的调查人员提供最先进的设备，以促进全面的评估方法 从亚细胞水平到整个动物。这些核心设施，围绕关键设备项目建造 设计通常比单个实验室提供的成本高，设计是由设计建立的 并推进我们直接影响退伍军人医疗保健的大型研究计划。 该核心提出的中心假设是提供高度敏感的先进技术 评估疾病或损伤的存在以及分子水平的进展，将增强生物学 我们的VA调查人员所做的工作的意义。该假设将通过4个初级解决 分析目标：1）对DNA和RNA靶标的高度敏感检测和绝对定量 1,000,000个总副本中有1个。 Bio-Rad的DDPCR非常适合研究低拷贝数成绩单 来自尺寸有限（啮齿动物组织等）的生物标本，或者存在于非常低的抽象性和 代表通常对于检测和分析人类疾病至关重要的靶标（即检查 循环生物标志物，如微RNA，圆形RNA，长的非编码RNA），可用于 诊断和预后； 2）在多功能和广泛使用的标准DNA和RNA分析技术 可扩展的工作流程（即基因分型，基因表达分析，拷贝数变化和下一代 测序验证）； 3）可再现的定量和蛋白质带上的蛋白质带（to） 使用标准化学发光方案和 高度敏感的成像系统，代替X射线膜； 4）多路复用荧光抗体检测 努力仅通过蛋白质印迹来识别多个分析物。这不仅减少了时间的时间， 而且还可以使用消耗品。鉴于我们核心设施的多功能性和综合性，VA 调查人员将能够从“每天”和高级技术协议中获取高质量的数据 可扩展的方式，以促进对血液，细胞和组织的生化分析作为关键成分 他们资助的研究。通过检查DNA，RNA和蛋白质获得的结果直接 适用于影响我们退伍军人的核心医学问题的病理生理学，例如血管疾病， 心脏病，神经系统疾病，肌肉骨骼病理学，中风，脑损伤和脊髓 受伤。提议的设备与我们现有的单元格和分子评估核心的集成将填充 技术差距，使我们的研究人员可以对其样品进行全方位服务的生化评估。这 代表一个独特的基础设施，将大大推动我们当地的研究 vamc。要求的仪器将进一步增强本地VA研究能力 在退伍军人事务部秘书中概述了卓越的蓝图蓝图。