Department of Biomedical Engineering, Ulsan National Institute of Science and Technology (UNIST), Ulsan 44919, Republic of Korea.
School of Materials Science and Engineering, UNIST, Ulsan 44919, Republic of Korea.
Biosens Bioelectron. 2019 Feb 1;126:36-43. doi: 10.1016/j.bios.2018.10.008. Epub 2018 Oct 19.
We present a novel vertical flow-based paper immunosensor for the rapid and sensitive electrochemical and colorimetric detection of influenza H1N1 viruses using a different pore size (DP) sample pad. The DP sample pad consisted of two different pore size papers: larger pores (diameter: 11 µm) facing the inlet, and smaller pores (diameter: 0.45 µm) facing the conjugate pad. This sample pad offered moderate and uniform flows, and hence concentrated horseradish peroxidase tagged antibodies (HRP-Abs)-H1N1 complexes from 40 µL of sample volumes on a conjugate pad for 2 min after sample injection, thereby providing fast detection (6 min for both detection methods) with 100 µL of flushing afterwards, high sensitivity, and the simplicity of the sensor. The filtration characteristics of the DP sample pad were evaluated using fluorescent beads, indicating that only small-sized bio-particles such as viruses can pass through the sample pad. Sandwich immunoreactions of HRP-Ab-H1N1-Ab were performed on the gold paper electrode of the immunoStrip, which was determined by electrochemical impedance spectroscopy (EIS) measurements. Simultaneously, the color signal of free HRP-Ab captured on the colorimetric zone was obtained using a scanner, and the intensity was analyzed using ImageJ. This immunosensor detected H1N1 virus concentration as low as 3.3 plaque forming units (PFU)/mL (phosphate buffer saline; PBS) and 4.7 PFU/mL (saliva) by EIS, and 1.34 PFU/mL (PBS) and 2.27 PFU/mL (saliva) by the colorimetric method. Furthermore, integrating these two detection methods can reduce false results with double assurance, and this device can provide a simple and economical on-site detection platform.
我们提出了一种新颖的基于垂直流动的纸基免疫传感器，用于快速灵敏地电化学和比色检测流感 H1N1 病毒，使用不同孔径（DP）的样品垫。DP 样品垫由两种不同孔径的纸张组成：较大的孔（直径：11 µm）朝向入口，较小的孔（直径：0.45 µm）朝向结合垫。这种样品垫提供了适度且均匀的流动，因此在样品注入后 2 分钟内，将来自 40 µL 样品体积的辣根过氧化物酶标记的抗体（HRP-Ab）-H1N1 复合物浓缩在结合垫上，从而提供快速检测（两种检测方法均为 6 分钟），之后用 100 µL 冲洗液冲洗，具有高灵敏度和传感器的简单性。使用荧光珠评估 DP 样品垫的过滤特性，表明只有小尺寸的生物颗粒（如病毒）才能通过样品垫。HRP-Ab-H1N1-Ab 的夹心免疫反应在免疫条的金纸电极上进行，通过电化学阻抗谱（EIS）测量来确定。同时，使用扫描仪获得固定在比色区的游离 HRP-Ab 的颜色信号，并使用 ImageJ 分析强度。该免疫传感器通过 EIS 检测到 H1N1 病毒浓度低至 3.3 噬菌斑形成单位（PFU）/mL（磷酸盐缓冲盐水；PBS）和 4.7 PFU/mL（唾液），通过比色法检测到 1.34 PFU/mL（PBS）和 2.27 PFU/mL（唾液）。此外，整合这两种检测方法可以通过双重保证减少假结果，并且该装置可以提供简单经济的现场检测平台。
