Utrasensitive bioassays using palladium catalyzed chemical amplification

描述（由申请人提供）：我们提议使用新型钯催化化学扩增（PdCCA）技术开发超灵敏生物测定法，该方法可检测血液中极低浓度的肿瘤生物标志物。PdCCA是一种通用技术，可普遍应用于不同的分析平台。在这里考虑的生物测定应用中，其使用用钯催化剂标记的检测抗体和随后的钯催化的高增益氧化还原反应，产生水溶性甲瓒染料。PdCCA将适用于标准ELISA形式，其中它可以在免疫测定中提供102至103倍的灵敏度增加因子，其中可获得高选择性的检测抗体。这种灵敏度的增加将降低目前使用的方法的检测阈值至ng-pg/ml的期望范围，从而允许开发用于早期肿瘤检测的筛选生物标志物。拟议工作的第一阶段将集中于技术开发和优化，使用一组卵巢癌的三种生物标志物和任选的乳腺癌生物标志物，目前正在华盛顿州西雅图的弗雷德哈钦森癌症研究中心（FHCRC）进行研究。我们将优化染料形成扩增化学和分光光度检测，并使用体外试验模型来开发和优化生物测定技术。我们的方法的可行性将在FHCRC储存的临床标本上得到证明。

DESCRIPTION (provided by applicant): We propose to use novel Palladium Catalyzed Chemical Amplification (PdCCA) technology to develop ultra-sensitive bioassays that can detect very low concentrations of tumor biomarkers in blood. PdCCA is a generic technology that can be universally applied to different assay platforms. In the bioassay application considered here it uses a detection antibody labeled with a palladium catalyst and a subsequent, palladium catalyzed, high gain redox reaction generating a water-soluble formazan dye. PdCCA will be adapted to the standard ELISA format, where it may provide a factor of 102 to 103 fold increase in sensitivity in immunoassays for which a detection antibody of high selectivity is available. This increase in sensitivity will lower the detection threshold of currently used methods to the desired range of ng-pg/ml, allowing for development of screening biomarkers for early tumor detection. Phase I of the proposed work will concentrate on technology development and optimization using a set of three biomarkers for ovarian cancer and, optionally, one biomarker of breast cancer, currently under study at the Fred Hutchinson Cancer Research Center, Seattle, WA (FHCRC). We will optimize the dye forming amplification chemistry and spectrophotometric detection, and use model in vitro assays to develop and optimize the bioassay technology. Feasibility of our approach will be demonstrated on clinical specimens stored at the FHCRC.