Scanning B. pseudomallei proteome for vaccine antigens

描述（由申请人提供）：由于潜在的生物恐怖主义威胁，开发一种安全有效的B。类鼻疽疫苗是国家和世界的目标。加州大学欧文分校的应用蛋白质组学实验室开发了一种名为PCR Express的高通量蛋白质表达系统，可用于从任何测序的微生物（包括A类、B类和C类生物恐怖主义制剂）快速生成完整的蛋白质组。该技术允许数百种不同的基因以每周数百种不同蛋白质的速度直接从其PCR产物中表达。在这里，将产生类鼻疽伯克霍尔德菌蛋白质组，并将其应用于识别B细胞和T细胞识别的抗原的问题，这些抗原将用于针对该试剂的亚单位疫苗。第一种形式的蛋白质组将在微阵列芯片上，该芯片将用于定量用合理减毒的B营养缺陷型接种的BALB/c小鼠的血清抗体滴度。类鼻疽（2D 2）和自然暴露于B的人。pseudomallei在流行地区的东北泰国对每一个单独的细菌蛋白。对于第二种蛋白质组形式，将纯化每种单独的蛋白质，并以使其能够用于体外T细胞再刺激试验的形式存在，该试验使用来自2D 2接种小鼠的脾细胞或来自暴露人群的全血。将通过ELISA试验测定lFN？分泌的T细胞活化。将在B感染的鼠模型中检测通过该疫苗抗原扫描鉴定的应答抗原的有效性。由于B.类鼻疽和B.鼻疽，在鼻疽动物模型中。这种定量的体液和细胞免疫应答扫描将产生针对B的免疫应答的第一个完整概况。在人类和实验动物中的类鼻疽杆菌，并将确定最有效的候选抗原，用于开发针对人类类鼻疽和鼻疽的DNA或亚单位疫苗。

DESCRIPTION (provided by applicant): Because of the potential bioterrorism threat, the development of a safe and effective B. pseudomallei vaccine is a national and worldwide goal. The Applied Proteomics Laboratory at UC Irvine has developed a high throughput protein expression system called PCR Express which can be used to rapidly generate complete proteomes from any sequenced microorganism, including the Class A, B & C Bioterrorism Agents. The technology allows hundreds of different genes to be expressed directly from their PCR products at the rate of hundreds of different proteins per week. Here the Burkholderia pseudomallei proteome will be generated and applied to the problem of identifying antigens recognized by B-cells and T-cells that will be useful in a subunit vaccine against the agent. The first form of the proteome will be on microarray chips which will be used to quantify serum antibody titers from BALB/c mice vaccinated with a rationally attenuated auxotroph of B. pseudomallei (2D2) and humans naturally exposed to B. pseudomallei in endemic regions of NE Thailand against each of the individual bacterial proteins. For the second proteome format, each individual protein will be purified and presented in a form that will enable them to be used in T cell restimulation assays in vitro using either spleen cells from 2D2 vaccinated mice or whole blood from exposed human populations. T cell activation will be determined by ELISA assay for secretion of lFN?. The responsive antigens identified by this Vaccine Antigen Scan will be tested for their efficacy in a murine model of infection with B. pseudomallei and, because of the extensive genetic similarity between B. pseudomallei and B. mallei, in animal models of glanders. This quantitative humoral and cellular immune response scan will produce the first complete profile of the immune response against B. pseudomallei in humans and experimental animals and will identify the most effective candidate antigens for development of a DNA or subunit vaccine against human melioidosis and glanders.