Large-scale Production & Purification Of Compounds With

大规模生产

• 批准号: 6983602
• 负责人: Joseph Shiloach
• 金额: --
• 依托单位: NATIONAL INSTITUTE OF DIABETES AND DIGESTIVE AND KIDNEY DISEASES
• 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/6983602
• 关键词: CHO cells; Corynebacterium diphtheriae; Escherichia coli; HeLa cells; Pseudomonas aeruginosa; Salmonella; Sf9 cell line; Shigella vaccines; bacterial proteins; biological products; biotechnology; chemical registry /resource; diphtheria toxin; fermentation; glucose metabolism; immunological substance; method development; microarray technology; microorganism mass culture; microorganism resource /registry; protein purification; tissue /cell culture; vaccine development

The Biotechnology Unit is responsible for the production and purification of biological material. The Unit undertakes a wide array of biological production tasks such as large scale growth of prokaryotes and eucaryotes, and production and purification of biological compounds especially proteins. These products, not available from commercial sources, are needed for clinical and structural studies. The Biotechnology Unit is capable of handling these various production tasks by implementing both physiological and technical approaches. Efficient process development and production is possible since the Unit possesses two integrated elements: a fully operational multi purpose pilot production facility and the capability of conducting research and process development. During the last year the unit performed approximately 200 different, large scale preparations including: 1) growing various microorganisms such as recombinant Escherichia coli, recombinant Pichia pastoris, mutant strains of Corynebacteria diphtheria, several types of Salmonella and Shigella, Staphylococcus strains and mutant strain of Bacillus anthracis; 2) propagating large volumes of various eucaryotic cells such as HeLa, CHO, drosophila, Sf 9 and hi five; 3) purifying gram quantities of diphtheria toxin CRM 9 (Cross Reacting Material), recombinant Pseudomonas aeruginosa exotoxin A, protective antigen from Bacillus anthracis. Staphylococcal enterotoxin type C, experimental transmission blocking vaccine for malaria and several antibodies both from hybridoma culture and rabbit serum. On the research and development arena, the unit continues its work in the following topics: 1) development of more efficient methods for large scale protein recovery, here we continue our work on implementing expanded bed adsorption techniques for capturing proteins on ion exchangers, and hydrophobic resins, utilizing new types of expanded column this column does not have flow adaptors and screens and therefore can handle larger concentration of biomass at higher low rates; 2) conducting research aimed at understanding the glucose metabolism of various recombinant E. coli strains in an effort to find out why certain strains are more suitable for foreign protein production using microarrays technology and protein analysis; 3) developing fermentation procedures for expressing sensitive, recombinant clostridium protein from E.coli by implementing anaerobic growth conditions; 4) conducting experiments for improving the production protocol of recombinant proteins from Pichia pastoris, especially experiment malaria transmission blocking vaccine; 5) the unit started another research project associated with mammalian cells, trying to understand the genetic difference between cells that can grow in suspension, and cells that are anchorage dependent. In addition, the unit continues its research work on the production process of recombinant proteins from the HeLa/vaccinia system, In collaboration with LDMI, NICHD, the unit conducted work on the development of new vaccine against Shigella flexneri and Bacillus anthracis, the products are currently in field trials.

生物技术股负责生物材料的生产和纯化。该股承担广泛的生物生产任务，如大规模生长原核生物和真核生物，以及生产和纯化生物化合物，特别是蛋白质。这些产品不能从商业来源获得，但需要用于临床和结构研究。生物技术股能够通过实施生理和技术方法来处理这些不同的生产任务。有效的工艺发展和生产是可能的，因为该股拥有两个综合要素：一个充分运作的多用途试验生产设施和进行研究和工艺发展的能力。去年，该单位进行了大约200种不同的大规模制备，包括：1)培养各种微生物，如重组大肠杆菌、重组毕赤酵母、白喉链杆菌突变菌株、几种沙门氏菌和志贺氏菌、葡萄球菌菌株和炭疽芽孢杆菌突变菌株；2)大量繁殖各种真核细胞，如HeLa、CHO、果蝇、Sf 9、hi 5等；3)纯化克量白喉毒素CRM 9（交叉反应物质）、重组铜绿假单胞菌外毒素A、炭疽芽孢杆菌保护性抗原。C型葡萄球菌肠毒素、实验性疟疾传播阻断疫苗及杂交瘤培养物和兔血清中几种抗体。在研究和开发领域，该部门继续开展以下主题的工作：1)开发更有效的大规模蛋白质回收方法，在这里，我们继续实施扩展床吸附技术，用于在离子交换器和疏水性树脂上捕获蛋白质，利用新型扩展柱，该柱没有流量适配器和筛网，因此可以以更高的低速率处理更高浓度的生物质；2)开展旨在了解各种重组大肠杆菌菌株葡萄糖代谢的研究，利用微阵列技术和蛋白质分析找出为什么某些菌株更适合外源蛋白生产；3)通过厌氧生长条件，开发从大肠杆菌中表达敏感、重组梭状芽胞杆菌蛋白的发酵工艺；4)开展改进毕赤酵母重组蛋白生产工艺的实验，特别是疟疾传播阻断疫苗实验；该单位开始了另一个与哺乳动物细胞相关的研究项目，试图了解可以在悬浮中生长的细胞和依赖于锚定的细胞之间的遗传差异。此外，该单位继续研究从HeLa/牛痘系统生产重组蛋白的过程。该单位与LDMI、NICHD合作，开展了针对福氏志贺氏菌和炭疽芽孢杆菌的新疫苗的开发工作，这些产品目前正在进行实地试验。