Fixed E. coli as Low Range Nucleic Acid Controls for M. tuberculosis Testing

固定大肠杆菌作为结核分枝杆菌检测的低范围核酸对照

• 批准号: 7254032
• 负责人: Clark A. Rundell
• 金额: $ 35.13万
• 依托单位: MAINE MOLECULAR QUALITY CONTROLS, INC.
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-06-15 至 2010-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7254032
• 关键词: Affect; Appendix; Bacillus (bacterium); Bacteria; Bacterial Counts; Behavior; Biological Assay; Biological Preservation; Bioterrorism; Buffers; Cell Wall; Cells; Cellular Structures; Cessation of life; Clinical; Clone Cells; Codon Nucleotides; Communicable Diseases; Condition; Cytolysis; DNA; DNA Sequence; Data; Decontamination; Depressed mood; Detection; Detergents; Development; Diagnosis; Diagnostic; Diagnostic tests; Digestion; Disease; Drug resistance; Electron Microscopy; Encapsulated; Ensure; Epidemic; Escherichia coli; Evaluation; Experimental Designs; Failure; Family; Fixatives; Future; Generations; Genes; Genome; Glutaral; Goals; Gordonia Bacterium; Health; Healthcare; Heating; Human Resources; Indium; Industry; Knowledge; Laboratories; Lead; Legal patent; Letters; Location; Medical; Membrane; Methods; Minor; Mission; Modification; Molecular; Molecular Diagnostic Techniques; Molecular Diagnostic Testing; Monitor; Multi-Drug Resistance; Multidrug-Resistant Tuberculosis; Mutate; Mutation; Mycobacterium bovis; Mycobacterium gordonae; Mycobacterium tuberculosis; National Institute of Allergy and Infectious Disease; New Jersey; None or Not Applicable; Nucleic Acid Amplification Tests; Nucleic Acids; Numbers; Outcome; Patients; Performance; Pharmaceutical Preparations; Phase; Plasmids; Polymerase Chain Reaction; Preparation; Prevention; Process; Product R; Property; Protocols documentation; Publications; Quality Control; Range; Reaction; Recovery; Regulation; Reporting; Research; Research Design; Research Infrastructure; Resistance; Ribosomal RNA; Sampling; Score; Signal Transduction; Simulate; Site; Small Business Funding Mechanisms; Small Business Innovation Research Grant; Source; Specificity; Specimen; Specimen Handling; Speed; Sputum; Technology; Test Result; Testing; Time; TimeLine; Tuberculosis; United States Dept. of Health and Human Services; Variant; Vial device; Work; base; cell fixing; commercial application; crosslink; desire; drug testing; gene cloning; improved; innovation; killings; medical schools; mutant; new technology; novel; nuclease; nucleic acid cloning; protein crosslink; prototype; quality assurance; research clinical testing; research study; respiratory; response; sample fixation; success; synthetic construct; tool; vpr Genes

DESCRIPTION (provided by applicant): The BROAD OBJECTIVE of this project is to produce a family of urgently needed quality controls for use in molecular Mycobacterium tuberculosis (M. tuberculosis) diagnostic tests. Quality controls are required by good laboratory practice and federal regulations to assure accuracy of all medical laboratory tests. The SPECIFIC AIMS are to 1) Produce E. Coli cells that parallel the properties of M. tuberculosis in DNA based molecular testing, 2) Clone gene sequences used to identify drug resistance in M. tuberculosis tests, and 3) Produce and evaluate prototype products. This Phase II work will develop a novel method to produce cloned E. coli with toughened, chemically crosslinked membranes containing synthetic M. tuberculosis DNA sequences for use as quality controls in molecular M. tuberculosis tests. Materials of this type are a necessary part of the infrastructure for the implementation of the new and rapid molecular technologies for M. tuberculosis testing. Efficient and accurate diagnoses, which are the promise of this new technology, are key to treatment and prevention of the major global health threat presented by M. tuberculosis. Enhancing the diagnostic tools and infrastructure enabling improved diagnostic testing is an important part of the NIAID mission. Further, in the "FOCUS on TB" agency mission: "NIAID supports a robust portfolio of research to develop new drugs and diagnostics----". The quality controls to be produced are a key element in developing and implementing new molecular diagnostic M. tuberculosis tests. In the RESEARCH DESIGN, molecular methods are used to produce clones that contain DNA sequences utilized in M. tuberculosis diagnosis. Mutant and wild type clones are constructed with sequences used in testing for drug resistance mutations. Methods traditionally used to fix membranes for electron microscopy studies are adapted to harden the membranes of the clone cells and to deactivate cellular nucleases. The fixing time, fixative adjuncts, and fixative concentrations are varied until the cell structure is hardened to a point where its release of nucleic acid during the clinical testing process is very similar to that of M. tuberculosis bacilli. The products will be evaluated in clinical laboratory molecular M. tuberculosis tests and fixing parameters adjusted to produce a material that is optimal for monitoring common molecular M. tuberculosis test.

描述（由申请人提供）：该项目的总体目标是生产一系列急需的质量控制品，用于分子结核分枝杆菌（M. tuberculosis）诊断测试。良好的实验室实践和联邦法规要求进行质量控制，以确保所有医学实验室测试的准确性。具体目标是 1) 在基于 DNA 的分子测试中产生与结核分枝杆菌特性相似的大肠杆菌细胞，2) 克隆用于在结核分枝杆菌测试中识别耐药性的基因序列，以及 3) 生产和评估原型产品。这项第二阶段工作将开发一种新方法来生产克隆大肠杆菌，该大肠杆菌具有含有合成结核分枝杆菌 DNA 序列的增韧化学交联膜，可用作分子结核分枝杆菌测试的质量控制。此类材料是实施结核分枝杆菌检测的新型快速分子技术的基础设施的必要组成部分。高效、准确的诊断是这项新技术的承诺，也是治疗和预防结核分枝杆菌带来的全球主要健康威胁的关键。增强诊断工具和基础设施以改进诊断测试是 NIAID 使命的重要组成部分。此外，在“关注结核病”机构使命中：“NIAID 支持强大的研究组合以开发新药和诊断方法——”。质量控制是开发和实施新的结核分枝杆菌分子诊断检测的关键要素。 在研究设计中，使用分子方法来产生含有用于结核分枝杆菌诊断的DNA序列的克隆。突变型和野生型克隆是用用于测试耐药性突变的序列构建的。传统上用于固定电子显微镜研究膜的方法适用于硬化克隆细胞的膜并使细胞核酸酶失活。改变固定时间、固定剂和固定剂浓度，直到细胞结构硬化到其在临床测试过程中释放的核酸与结核分枝杆菌非常相似的程度。这些产品将在临床实验室分子结核分枝杆菌测试中进行评估，并调整参数以生产最适合监测常见分子结核分枝杆菌测试的材料。