Division of Solid-State Electronics, Department of Electrical Engineering, Uppsala University, 75 121 Uppsala, Sweden.
Department of Applied Physics, School of Engineering Sciences, KTH Royal Institute of Technology, 10 691 Stockholm, Sweden.
ACS Sens. 2024 Jun 28;9(6):2935-2945. doi: 10.1021/acssensors.4c00110. Epub 2024 Jun 7.
Detection of analytes using streaming current has previously been explored using both experimental approaches and theoretical analyses of such data. However, further developments are needed for establishing a viable microchip that can be exploited to deliver a sensitive, robust, and scalable biosensor device. In this study, we demonstrated the fabrication of such a device on silicon wafer using a scalable silicon microfabrication technology followed by characterization and optimization of this sensor for detection of small extracellular vesicles (sEVs) with sizes in the range of 30 to 200 nm, as determined by nanoparticle tracking analyses. We showed that the sensitivity of the devices, assessed by a common protein-ligand pair and sEVs, significantly outperforms previous approaches using the same principle. Two versions of the microchips, denoted as enclosed and removable-top microchips, were developed and compared, aiming to discern the importance of high-pressure measurement versus easier and better surface preparation capacity. A custom-built chip manifold allowing easy interfacing with standard microfluidic connections was also constructed. By investigating different electrical, fluidic, morphological, and fluorescence measurements, we show that while the enclosed microchip with its robust glass-silicon bonding can withstand higher pressure and thus generate higher streaming current, the removable-top configuration offers several practical benefits, including easy surface preparation, uniform probe conjugation, and improvement in the limit of detection (LoD). We further compared two common surface functionalization strategies and showed that the developed microchip can achieve both high sensitivity for membrane protein profiling and low LoD for detection of sEV detection. At the optimum working condition, we demonstrated that the microchip could detect sEVs reaching an LoD of 10 sEVs/mL (when captured by membrane-sensing peptide (MSP) probes), which is among the lowest in the so far reported microchip-based methods.
使用流动电流检测分析物的方法之前已经通过实验方法和对这些数据的理论分析进行了探索。然而，需要进一步的发展来建立一种可行的微芯片，可以用来提供灵敏、稳健和可扩展的生物传感器设备。在这项研究中，我们使用可扩展的硅微制造技术在硅片上展示了这种设备的制造，然后对该传感器进行了表征和优化，以检测纳米颗粒跟踪分析确定的尺寸在 30 至 200nm 范围内的小细胞外囊泡 (sEV)。我们表明，通过常见的蛋白质-配体对和 sEV 评估的器件灵敏度明显优于使用相同原理的先前方法。开发了两种微芯片版本，分别称为封闭和可移除顶微芯片，并进行了比较，旨在区分高压测量与更容易和更好的表面准备能力的重要性。还构建了一个定制的芯片歧管，允许与标准微流控连接轻松接口。通过研究不同的电气、流体、形态和荧光测量，我们表明，虽然具有坚固的玻璃-硅键合的封闭微芯片可以承受更高的压力，从而产生更高的流动电流，但可移除顶配置提供了几个实际优势，包括易于表面准备、均匀的探针缀合以及检测限 (LoD) 的提高。我们进一步比较了两种常见的表面功能化策略，并表明开发的微芯片可以实现高灵敏度的膜蛋白分析和低 LoD 的 sEV 检测。在最佳工作条件下，我们证明微芯片可以检测到 sEV，达到 10sEV/mL 的 LoD（当被膜感应肽 (MSP) 探针捕获时），这在迄今为止报道的基于微芯片的方法中是最低的之一。
