Phage Display Tools for Automated Blood Typing

用于自动血型定型的噬菌体展示工具

• 批准号: 7254809
• 负责人: DONALD Lawrence SIEGEL
• 金额: $ 50.95万
• 依托单位: PHENOTECH, INC.
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-05-01 至 2009-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7254809
• 关键词: Address; Agglutination; Alloimmunization; Amino Acid Sequence; Antibodies; Antigen Receptors; Antigens; Automation; Bacteriophages; Biological Assay; Blood; Blood Banks; Blood Group Antigens; Blood Tests; Blood typing procedure; Cells; Class; Clinical; Collection; Compatible; Complement; Complement 3d; Coombs' Test; Coupled; DNA; DNA Sequence; Detection; Development; Ensure; Erythrocytes; Funding; Future; Genotype; Globulins; Human; Immunoglobulin G; Immunoglobulin M; Incidence; Inovirus; Laboratories; Length; Link; MNSs Blood-Group System; Manuals; Materials Testing; Medical; Medicine; Methodology; Methods; Microfluidics; Molecular; Monoclonal Antibodies; Mus; Nucleic Acids; Numbers; Oryctolagus cuniculus; Performance; Phage Display; Phase; Phase I Clinical Trials; Phenotype; Plasma; Reaction; Reagent; Reporting; Sampling; Scheme; Sensitivity and Specificity; Services; Specificity; Surface; System; Technology; Testing; Time; Transfusion; Tube; United States Food and Drug Administration; Work; antigen antibody binding; base; cell type; cost; improved; instrumentation; micro-total analysis system; novel; nucleic acid detection; particle; programs; reagent testing; receptor; tool

DESCRIPTION (provided by applicant): Current technologies used in blood banking are extraordinarily labor intensive, prone to human error, and an order of magnitude more expensive per test that those in other clinical laboratories. Coupled with a growing shortage of skilled medical technologists, dwindling supplies of human plasma-derived phenotyping reagents, and an inherent difficulty in fully automating agglutination-based methodologies, the ability to perform rapid and accurate pre-transfusion testing in a cost-effective manner has become a significant challenge. PhenoTech's long-term objective is to use a set of novel molecular technologies to develop a new class of renewable, inexpensive, high-quality blood bank testing reagents that will function in a rapid, high- throughput, automatable assay system. At the core of the proposed technology are red blood cell antigen- specific monoclonal antibodies displayed on the surface of bacteriophage particles. Phase I studies proposed to exploit the naturally-occurring presence of unique DNA sequences within the particles to develop an assay system in which the phenotype of a cell is determined by assaying the genotype of the detecting reagent. Such a strategy would offer extraordinary sensitivity and specificity, would require minute amounts of testing materials and reagents, would be easily adapted to automation, and would be amenable to multiplexing strategies offering the possibility of simultaneous antigen profiling of a red cell sample in a single reaction vessel. This would make extended phenotyping and matching of blood beyond the traditional three-antigen A, B, and Rh(D) affordable and routine. Such a paradigm shift in the practice of transfusion medicine would then serve to reduce the incidence of red cell alloimmunization and occurrence of delayed hemolytic transfusion reactions, decrease the development of positive antibody screens and the costs in working them up, and improve the turn-around-time for the provision of immunologically-compatible blood. As reported in this Phase II proposal, PhenpTech was successful in demonstrating the feasibility of multiplexing red cell typing reactions using its novel reagent genotyping methodology. Reactions required less than 1/100,000 the number of RBCs of a conventional agglutination assay and used attogram equivalents of conventional antibody reagents. In addition, the ability to use phage particles which express anti-globulin-like molecules for performing indirect antiglobulin tests, the most labor-intensive and costly group of pre-transfusion tests, was also shown. Having demonstrated the feasibility of phage reagent genotyping methods, PhenoTech's Phase II program will now focus on the development of a suite of phage reagents to clinically important blood group antigens required for further development of its blood testing technologies. Combined with its concurrent development of a microfluidics-based "lab-on-a-chip" platform, PhenoTech will be able to provide the tools necessary to ensure the safe practice of transfusion medicine in the future by addressing the technical and fiscal limitations of conventional blood bank testing methods.

描述（由申请人提供）：血液库中使用的当前技术非常密集，容易出现人为错误，并且每次测试的数量级比其他临床实验室中的测试昂贵。再加上熟练的医疗技术人员的越来越短，人类等离子体衍生的表型试剂的供应减少，以及完全自动化基于凝集的方法论的固有困难，以成本效益的方式进行快速，准确的预交易前输血测试的能力已成为一项重大挑战。 Phanotech的长期目标是使用一组新型的分子技术来开发一类新的可再生，廉价，高质量的血库测试试剂，这些试剂将在快速，高吞吐量，可自动性测定系统中起作用。所提出的技术的核心是噬菌体颗粒表面上显示的红细胞抗原特异性单克隆抗体。第一阶段的研究提议利用颗粒中独特的DNA序列的自然存在，以开发一个测定系统，在该系统中，通过分析检测试剂的基因型来确定细胞的表型。这种策略将提供非凡的敏感性和特异性，需要微量的测试材料和试剂，很容易适应自动化，并且可以使单个反应容器中红细胞样品同时抗原分析的可能性多重策略。这将使血液的扩展表型和匹配超出传统的三抗A，B和RH（D）负担得起的和常规的效果。然后，输血医学实践的这种范式转移将有助于减少红细胞同种异体免疫的发生率，并发生延迟的溶血输血反应，减少阳性抗体筛查的发展以及工作成本的发展，并提高其转折时间，以提供免疫兼容的血液。正如这一II期提案所报道的那样，Phenptech成功地证明了使用其新型试剂基因分型方法多路复用红细胞键入反应的可行性。反应需要少于1/100,000的常规凝集测定法的RBC数量，并使用了常规抗体试剂的图表等效物。此外，还显示了使用表达抗全球蛋白样分子进行间接抗球蛋白测试的噬菌体颗粒的能力，这是最富有劳动力且昂贵的转交前输血测试组。在证明了噬菌体试剂基因分型方法的可行性之后，Pachotech的II期计划现在将着重于开发一套噬菌体试剂，以进一步开发其血液测试技术所需的临床重要血型抗原。结合其同时开发基于微流体的“芯片实验室”平台，Phenotech将能够通过解决常规血库测试方法的技术和财政限制，提供必要的工具，以确保将来的输血医学实践安全。