TITER ON CHIP: ALTERNATIVE TO SRID FOR INFLUENZA VACCINE POTENCY DETERMINATION

描述（由申请人提供）：迫切需要新的分析方法来加强疫苗的研究和开发，最终能够在更短的时间内以更低的成本生产安全有效的疫苗。例如，众所周知，对于分裂疫苗来说，过程中可能限制速率的一个步骤是蛋白质定量和效力测定。FDA批准的流感血凝素蛋白疫苗效价测定的“金标准”是单径向免疫扩散（SRID）。SRID是一种时间和劳动密集型分析，通常需要2-3天才能完成，训练有素的分析人员至少需要6小时的时间。虽然参考试剂由生物制品评估和研究中心（CBER）免费提供，但必须购买额外的材料，并由每个疫苗生产商准备和验证整个分析。疫苗生产商通常在收到参考试剂方面经历长时间的延误，有时长达数月。即使有参考资料在手，在进行开发之前，每一轮克隆评估的等待结果可能需要几天时间。正如人们普遍承认的那样，总体结果是一个耗时和低效的疫苗开发过程。本文提出了两种新的基于低成本低密度微阵列的定量、多路分析方法。这两种检测方法都基于“芯片滴度”方法，该方法将通过大大缩短获得结果的时间、消除与检测制剂相关的实验室间差异和降低试剂成本来简化疫苗效力测量。一种被提议的检测方法依赖于（Specific Aim I）对流感血凝素亚型（如H1、H3、H5）普遍反应的单克隆抗体。另一种被提议的检测方法（Specific Aim II）依靠与血凝素结合的通用唾液酸糖蛋白来实现快速的血凝素蛋白定量，而不需要菌株特异性抗体。我们相信，滴度芯片具有革命性的流感疫苗效力测定的长期潜力。

DESCRIPTION (provided by applicant): There is a tremendous need for new analytical methods to enhance vaccine research and development, ultimately allowing the production of safe and efficacious vaccines in less time at less cost. For example, it is well known that for splt vaccines one step in the process that can be rate limiting is protein quantification and potency determination. The FDA approved "gold standard" potency assay for influenza hemagglutinin protein based vaccines is single radial immunodiffusion (SRID). SRID is a time and labor intensive assay, often requiring 2-3 days to complete and a minimum of 6 hours hands on time by well trained analysts. While the reference reagents are provided at no cost by the Center for Biologics Evaluation and Research (CBER), additional materials must be purchased and the entire assay prepared and validated by each vaccine producer. Often vaccine producers experience long delays, sometime months, in receiving reference reagents. Even with reference materials in hand, the wait for results can be days for each round of clone assessment prior to moving forward in development. As is widely acknowledged, the overall result is a time-consuming and inefficient vaccine development process. Here we propose two new quantitative, multiplexed analytical methods based on cost-effective low density microarrays. Both assays are based on a ¿Titer on Chip¿ approach that will streamline vaccine potency measurements by substantially reducing time to result, eliminating inter-laboratory variations associated with assay preparations, and reducing reagent cost. One proposed assay relies (Specific Aim I) on monoclonal antibodies that are universally responsive to hemagglutinin subtypes for influenza (e.g., H1, H3, H5). The other proposed assay (Specific Aim II) relies on universal sialic acid glycoproteins that bind hemagglutinin to achieve rapid HA protein quantification without the need for strain specific antibodies. We believe that Titer on Chip has the long term potential to revolutionize influenza vaccine potency determination. PUBLIC HEALTH RELEVANCE: There is a tremendous need for new analytical methods to enhance vaccine research and development, ultimately allowing the production of safe and efficacious vaccines in less time at less cost. Here we propose two new quantitative, multiplexed analytical methods based on cost-effective low density microarrays. We believe that Titer on Chip has the long term potential to revolutionize influenza vaccine potency determination.