NISED - A Nanoparticle Immunoassay based on Single-entity Electrochemical Detection

本提案的目的是研究一种新的分析技术，用于快速和选择性定量的生物分子，使用单实体电化学检测芯片。我们将利用巯基连接的银纳米颗粒与识别分子，包括抗体和适配体，用于特异性结合溶液中的靶分子。传感器系统将包括用于捕获和检测纳米颗粒的紧密间隔且可单独寻址的电极的阵列。一些电极将通过硫醇-金相互作用用识别分子官能化，以在与分析物的初步孵育阶段期间用作靶分子的捕获探针。在孵育和暴露于官能化纳米颗粒之后，将还原脉冲施加到捕获电极以通过破坏金属-硫键来释放结合的纳米颗粒。释放的颗粒随后将在相邻的检测电极处经由冲击电化学检测。在此过程中，银纳米颗粒将被氧化，产生短且限定的电流尖峰。由于纳米颗粒碰撞的频率取决于颗粒到检测电极的质量传输，因此记录的电流尖峰的速率将反映捕获的纳米颗粒的数量和分析物中靶分子的浓度。微流体限制将确保纳米颗粒的检测效率接近100%。总的来说，所提出的概念将提供一种基于单实体纳米颗粒检测的具有电子读数的高灵敏度免疫测定。作为概念验证，我们将研究该检测试剂盒选择性检测血清和全血中胰岛素的性能。

The aim of this proposal is to investigate a new analytical technique for rapid and selective quantification of biomolecules using single-entity electrochemical detection on a chip. We will make use of thiol-linked silver nanoparticles with recognition molecules including antibodies and aptamers for specifically binding target molecules in solution. The sensor system will comprise an array of closely spaced and individually addressable electrodes for capturing and detecting nanoparticles. Some of the electrodes will be functionalized with recognition molecules via thiol-gold interactions to serve as capture probes for the target molecules during a preliminary incubation phase with the analyte. After incubation and exposure to functionalized nanoparticles, a reductive pulse will be applied to the capture electrodes to release bound nanoparticles by breaking the metal-sulfur bonds. The released particles will subsequently be detected at adjacent detection electrodes via impact electrochemistry. During this process, the silver nanoparticles will be oxidized generating short and defined current spikes. As the frequency of nanoparticle impacts depends on the mass transport of particles to the detection electrode, the rate of recorded current spikes will reflect the number of captured nanoparticles and the concentration of the target molecules in the analyte. A microfluidic confinement will ensure a detection efficiency of the nanoparticles close to 100%. Overall, the proposed concept will provide a highly sensitive immunoassay with electronic readout based on single-entity nanoparticle detection. As a proof of concept we will investigate the performance of this assay for the selective detection of insulin in serum as well as whole blood.