Rapid homogeneous antibody-based detection of proteins

描述（由申请人提供）：各种生物样品中蛋白质的检测和定量是最重要和最常用的分析方法之一，应用于研究，医学诊断，早期疾病检测和生物威胁剂检测。虽然有许多可用的蛋白质检测方法（ELISA是其中的金标准），但它们都有一些局限性。最常见的限制是进行分析所需的时间和分析中涉及的样品操作的复杂性。因此，显然需要开发新的快速和均质蛋白质检测方法。在该项目的第一阶段，我们建立了一种新的基于抗体的均质蛋白检测方法（分子钳子）的可行性，该方法允许通过简单的荧光强度测量来检测目标蛋白的存在。该分析快速，技术上操作极其简单（除了将样品与检测试剂混合外，基本上不涉及样品操作）。这些特点使得这种新的检测格式在没有配备精密仪器的实验室支持而必须快速确定目标蛋白的情况下特别有用（例如在即时诊断检测的情况下）。该项目的长期目标是利用分子钳开发简单、快速和廉价的即时诊断测试，用于一组心脏病标志物。该二期项目的目标是为研究市场开发六种心脏病标志物（肌钙蛋白I、b型利钠肽（BNP）、肌红蛋白、肌酸激酶（CK-MB）、c反应蛋白（CRP）和d -二聚体）的分子钳法。这一目标的完成将允许启动批准这些检测用于临床诊断的程序。此外，我们将通过制定提高检测灵敏度的策略来加强分子钳检测。这一目标的完成将扩大分子钳的可能靶标范围，从而提高分子钳的商业化前景。

DESCRIPTION (provided by applicant): Detection and quantification of proteins in various biological samples is one of the most important and most often utilized assays with applications in research, medical diagnosis, early disease detection and detection of biological threat agents. While many protein detection methodologies are available (ELISA being a gold standard of those), they each have some limitations. The most common limitation is the time required to perform the assay and the complexity of the sample manipulation involved in the assay. Thus, there is a clear need for developing new rapid and homogenous protein detection methodologies. In Phase I of this project we established feasibility of a novel antibody-based homogenous protein detection methodology (molecular pincers) which allows detection of the presence of a target protein by a simple fluorescence intensity measurement. The assay is rapid and is technically extremely simple to perform (involves essentially no sample manipulation other then mixing the sample with the detection reagent). These characteristics make this new assay format particularly useful in situations where quick determination of the target protein is essential without a support of a laboratory equipped with sophisticated instrumentation (as in the case, for example, of point-of-care diagnostic assay). The long-term goal of this project will be to utilize molecular pincers to develop simple, rapid and inexpensive point-of-care diagnostic tests for a panel of cardiac disease markers. The goal of this Phase II project will be to develop molecular pincer assays for six markers of cardiac disease (troponin I, B-type natriuretic peptide (BNP), myoglobin, creatine kinase (CK-MB), C-reactive protein (CRP), and D-dimer) for research market. Completion of this goal will allow initiation of the process of approval of these assays for clinical diagnosis. Additionally, we will work on enhancing molecular pincer assay by developing strategies to improve assay sensitivity. Completion of this goal will enhance the commercialization prospects of molecular pincers by broadening the range of possible targets for which the assay could be developed. PUBLIC HEALTH RELEVANCE: Molecular pincer assays will find wide applications in research and diagnosis of human disease. In research, it could become more convenient, easier to use, less expensive replacement for currently used assays for detecting proteins. In diagnosis, it will allow more rapid and more straightforward detection of antigens correlated with human disease.