Department of Chemistry, Khalifa University of Science and Technology, Abu Dhabi, P.O. Box 127788, United Arab Emirates.
Department of Chemistry, Khalifa University of Science and Technology, Abu Dhabi, P.O. Box 127788, United Arab Emirates; Center for Catalysis and Separations, Khalifa University of Science and Technology, Abu Dhabi, P.O. Box 127788, United Arab Emirates.
Biosens Bioelectron. 2024 Sep 1;259:116388. doi: 10.1016/j.bios.2024.116388. Epub 2024 May 15.
Claudin18.2 (CLDN18.2) is a tight junction protein often overexpressed in various solid tumors, including gastrointestinal and esophageal cancers, serving as a promising target and potential biomarker for tumor diagnosis, treatment assessment, and prognosis. Despite its significance, no biosensor has been reported to date for the detection of CLDN18.2. Here, we present the inaugural immunosensor for CLDN18.2. In this study, an amine-rich conducting polymer of polymelamine (PM) was electrografted onto different carbon nanomaterial-based screen-printed electrodes (SPEs), including carbon (C), graphene (Gr), graphene oxide (GO), carbon nanotube (CNT), and carbon nanofiber (CNF) via cyclic voltammetry. A comparative study was performed to explore the best material for the preparation of the PM-modified electrodes to be used as in-situ redox substrate for the immunosensor fabrication. The surface chemistry and structural features of pristine and PM-deposited electrodes were analyzed using Raman and scanning electron microscopy (SEM) techniques. Our results showed that the PM deposited on Gr and CNT/SPEs exhibited the most significant and stable redox behavior in PBS buffer. The terminal amine moieties on the PM-modified electrode surfaces were utilized for immobilizing anti-CLDN18.2 monoclonal antibodies via N-ethyl-N'-(3-(dimethylamino)propyl)carbodiimide/N-hydroxysuccinimide chemistry to construct the electrochemical immunosensor platform. Differential pulse voltammetry-based immunosensing of CLDN18.2 protein on BSA/anti-CLDN18.2/PM-Gr/SPE and BSA/anti-CLDN18.2/PM-CNT/SPE exhibited excellent selectivity against other proteins such as CD1, PDCD1, and ErBb2. The limits of detection of these two immunosensor platforms were calculated to be 7.9 pg/mL and 0.104 ng/mL for the CNT and Gr immunosensors, respectively. This study demonstrated that the PM-modified Gr and CNT electrodes offer promising platforms not only for the reagentless signaling but also for covalent immobilization of biomolecules. Moreover, these platforms offer excellent sensitivity and selectivity for the detection of CLDN18.2 due to its enhanced stable redox activity. The immunosensor demonstrated promising results for the sensitive detection of CLDN18.2 in biological samples, addressing the critical need for early gastric cancer diagnosis.
Claudin18.2（CLDN18.2）是一种紧密连接蛋白，在包括胃肠道和食管癌在内的各种实体瘤中常过度表达，是肿瘤诊断、治疗评估和预后的有前途的靶标和潜在生物标志物。尽管它很重要，但迄今为止还没有报道用于检测 CLDN18.2 的生物传感器。在这里，我们提出了首个用于 CLDN18.2 的免疫传感器。在这项研究中，通过循环伏安法将富含胺的三聚氰胺聚合物（PM）电接到不同的基于碳纳米材料的丝网印刷电极（SPE）上，包括碳（C）、石墨烯（Gr）、氧化石墨烯（GO）、碳纳米管（CNT）和碳纳米纤维（CNF）。进行了一项比较研究，以探索制备 PM 修饰电极的最佳材料，该电极可用作免疫传感器制造的原位氧化还原底物。使用拉曼和扫描电子显微镜（SEM）技术分析了原始和 PM 沉积电极的表面化学和结构特征。我们的结果表明，在 PBS 缓冲液中，沉积在 Gr 和 CNT/SPE 上的 PM 表现出最显著和稳定的氧化还原行为。PM 修饰电极表面上的末端胺基用于通过 N-乙基-N'-(3-(二甲氨基)丙基)碳二亚胺/N-羟基琥珀酰亚胺化学固定抗 CLDN18.2 单克隆抗体，构建电化学免疫传感器平台。基于差分脉冲伏安法的 BSA/抗 CLDN18.2/PM-Gr/SPE 和 BSA/抗 CLDN18.2/PM-CNT/SPE 上 CLDN18.2 蛋白的免疫传感显示出对其他蛋白质（如 CD1、PDCD1 和 ErBb2）的出色选择性。这两个免疫传感器平台的检测限分别计算为 CNT 和 Gr 免疫传感器的 7.9 pg/mL 和 0.104 ng/mL。这项研究表明，PM 修饰的 Gr 和 CNT 电极不仅为无试剂信号提供了有前途的平台，而且为生物分子的共价固定提供了平台。此外，由于其增强的稳定氧化还原活性，这些平台对 CLDN18.2 的检测具有出色的灵敏度和选择性。该免疫传感器在生物样品中对 CLDN18.2 的灵敏检测表现出有前景的结果，满足了早期胃癌诊断的关键需求。
