Department of Materials and Environmental Technology, Tallinn University of Technology, Ehitajate tee 5, 19086, Tallinn, Estonia.
Russian Academy of Sciences, Institute of Macromolecular Compounds, Bolshoy prospect 31, 199004, St.Petersburg, Russia.
Anal Biochem. 2021 Jul 1;624:114196. doi: 10.1016/j.ab.2021.114196. Epub 2021 Apr 15.
This paper is aimed at the development of a biosensor for direct detection of Hepatitis C virus (HCV) surface antigen: envelope protein (E2). A recombinant LEL fragment of biological cell receptor CD81 and two short synthetic peptides imitating the fragment of LEL sequence of CD81 (linear and loop-like peptides) capable of specific binding to E2 were tested as molecular recognition elements of the biosensor. For this purpose the selected ligands were immobilized to the surface of a screen-printed electrode utilized as an electrochemical sensor platform. The immobilization parameters such as the ligand concentration and the immobilization time were carefully optimized for each ligand. Differential pulse voltammetry used to evaluate quantitatively binding of E2 to the ligands revealed their similar binding affinity towards E2. Thus, the linear peptide was selected as a less expensive and easily prepared ligand for the HCV biosensor preparation. The resulting HCV biosensor demonstrated selectivity towards E2 in the presence of interfering protein, conalbumin. Moreover, it was found that the prepared biosensor effectively detected E2 bound to hepatitis C virus-mimetic particles (HC VMPs) at LOD value of 2.1∙10 mg/mL both in 0.01 M PBS solution (pH 7.4) and in simulated blood plasma.
本文旨在开发一种用于直接检测丙型肝炎病毒 (HCV) 表面抗原：包膜蛋白 (E2) 的生物传感器。作为生物传感器的分子识别元件，我们测试了生物细胞受体 CD81 的重组 LEL 片段和两个模仿 CD81 的 LEL 序列片段（线性和环肽）的短合成肽，这两种肽能够与 E2 特异性结合。为此，选择的配体被固定在用作电化学传感器平台的丝网印刷电极的表面上。仔细优化了每种配体的配体浓度和固定化时间等固定化参数。用于定量评估 E2 与配体结合的差分脉冲伏安法显示它们对 E2 具有相似的结合亲和力。因此，选择线性肽作为用于 HCV 生物传感器制备的更便宜且易于制备的配体。所制备的 HCV 生物传感器在存在干扰蛋白卵清白蛋白的情况下对 E2 表现出选择性。此外，还发现制备的生物传感器在 0.01 M PBS 溶液（pH 7.4）和模拟血浆中，在 HCV 模拟颗粒（HCVMPs）上检测到 E2 的检测限值（LOD）为 2.1×10-3mg/mL。
