Immunoproteomics and detection of viral diseases

描述(由申请人提供)：传染病病原体的快速检测与检测生物恐怖分子病原体的问题变得更加相关。目前临床相关分子的检测和定量技术是基于用于免疫分析或使用RT-PCR系统检测核酸的单克隆和多克隆抗体。这些检测相对较慢，至少需要3个小时才能完成。提出了一种新的、快速的、基于高亲和力特异性抗体与质谱读出相结合的免疫蛋白质组学分析方法。在一系列初步研究中，我们证明了利用表面增强激光解吸电离飞行时间质谱仪(SELDI-TOF)，使用单一抗体包被的抗原捕获和转移试剂(ACTR)以良好的灵敏度检测模式抗原的能力。这种免疫蛋白质组学分析可以在1小时内完成，并遵循如下顺序过程：在步骤1中固定抗体，在步骤2中捕获抗原，在步骤3中通过质谱分析结合的抗原。免疫蛋白质组分析可能比目前的抗体分析更快和敏感，并可以为感染动物的样本中细菌或病毒抗原或针对这些产品的特定抗体提供证据。这份R03提案旨在优化这一平台技术。在最初的概念验证研究中，我们将开发和评估免疫蛋白质组分析方法，以测量感染癫痫小鼠血清中的干扰素-γ(Th1细胞因子)和特异性抗病毒抗体。皮疹是CDC A类生物恐怖分子天花病毒的替代品，因此，成功的免疫蛋白质组学方法可能与生物恐怖分子的检测有关。

DESCRIPTION (provided by applicant): Rapid detection of infectious agents has become more relevant with concerns associated with detection of bioterrorist agents. Current technology for detection and quantification of clinically relevant molecules is based on monoclonal and polyclonal antibodies used for immunoassay or detection of nucleic acids using RT-PCR systems. These assays are relatively slow taking at least 3 hours to complete. A novel, rapid, immunoproteomics assay based on high affinity specific antibodies coupled with a mass spectral read out is proposed. In a series of preliminary studies we have demonstrated the ability to use a single antibody-coated antigen capture and transfer reagent (ACTR) to detect model antigens, with good sensitivity, using surface enhanced laser desorption ionization time-of-flight mass spectrometry (SELDI-TOF). This immunoproteomic assay can be complete in < 1 hour and follows a sequential process in which antibody is immobilized in step 1, antigen is captured in step 2 and bound antigen is analyzed by mass spectrometry in step 3. Immunoproteomic assays may potentially be faster and as sensitive as the current antibody assays and could provide evidence for bacterial or viral antigens or specific antibodies to these products in samples for an infected animal. This RO3 proposal is designed to optimize this platform technology. In the initial proof-of-concept studies, we will develop and evaluate immunoproteomic assays to measure INF gamma (Th1 cytokine) and specific anti-viral antibodies in the serum of ectromelia infected mice. Ectromelia is a surrogate of variola virus, a CDC Category A bioterrorist agent and thus, the successful immunoproteomic approach may be relevant for detection of bioterrorist agents.