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，用于快速和选择性地定量生物分子，使用芯片上的单实体电化学检测。我们将利用巯基银纳米颗粒与识别分子（包括抗体和适配体）特异性结合溶液中的靶分子。该传感器系统将包括一组间距紧密且可单独寻址的电极，用于捕获和检测纳米颗粒。一些电极将通过巯基-金相互作用与识别分子功能化，在与分析物的初步孵育阶段作为目标分子的捕获探针。在孵育和暴露于功能化纳米粒子后，将在捕获电极上施加还原脉冲，通过破坏金属-硫键来释放结合的纳米粒子。释放的颗粒随后将通过冲击电化学在相邻的检测电极上检测到。在这个过程中，银纳米粒子将被氧化，产生短而明确的电流尖峰。由于纳米粒子撞击的频率取决于粒子到检测电极的质量传递，记录的电流峰值的速率将反映捕获的纳米粒子的数量和分析物中目标分子的浓度。微流体约束将确保纳米颗粒的检测效率接近100%。总的来说，提出的概念将提供一种基于单实体纳米颗粒检测的高灵敏度电子读数免疫分析。作为一个概念的证明，我们将研究这种方法的性能选择性检测胰岛素在血清和全血。

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.