State Key Laboratory of Chemo/Bio-Sensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha, 410082, P R China.
Nucleic Acids Res. 2011 Oct;39(18):e122. doi: 10.1093/nar/gkr559. Epub 2011 Jul 8.
Development of novel aptamer sensor strategies for rapid and selective assays of protein biomarkers plays crucial roles in proteomics and clinical diagnostics. Herein, we have developed a novel aptamer sensor strategy for homogeneous detection of protein targets based on fluorescence protection assay. This strategy is based on our reasoning that interaction of aptamer with its protein target may dramatically increase steric hindrance, which protects the fluorophore, fluorescein isothiocyannate (FITC), labeled at the binding pocket from accessing and quenching by the FITC antibody. The aptamer sensor strategy is demonstrated using a model protein target of immunoglobulin E (IgE), a known biomarker associated with atopic allergic diseases. The results reveal that the aptamer sensor shows substantial (>6-fold) fluorescence enhancement in response to the protein target, thereby verifying the mechanism of fluorescence protection. Moreover, the aptamer sensor displays improved specificity to other co-existing proteins and a desirable dynamic range within the IgE concentration from 0.1 to 50 nM with a readily achieved detection limit of 0.1 nM. Because of great robustness, easy operation and scalability for parallel assays, the developed homogeneous fluorescence protection assay strategy might create a new methodology for developing aptamer sensors in sensitive, selective detection of proteins.
开发新型适体传感器策略，用于快速和选择性地分析蛋白质生物标志物，在蛋白质组学和临床诊断中发挥着至关重要的作用。本文开发了一种基于荧光保护分析的新型适体传感器策略，用于均相检测蛋白质靶标。该策略基于我们的推理，即适体与蛋白质靶标的相互作用可能会显著增加空间位阻，从而保护结合口袋处标记的荧光染料异硫氰酸荧光素（FITC）免受 FITC 抗体的接近和淬灭。该适体传感器策略使用免疫球蛋白 E（IgE）作为模型蛋白靶标进行了验证，IgE 是与特应性过敏疾病相关的已知生物标志物。结果表明，该适体传感器对蛋白质靶标表现出显著的（>6 倍）荧光增强，从而验证了荧光保护的机制。此外，该适体传感器对其他共存蛋白具有改善的特异性，并且在 IgE 浓度为 0.1 至 50 nM 的范围内具有理想的动态范围，检测限低至 0.1 nM。由于具有强大的稳健性、易于操作和可扩展性，用于平行分析，所开发的均相荧光保护分析策略可能为基于适体的蛋白质敏感、选择性检测开发新型传感器提供了一种新方法。
