Department of Medicinal Plant Production, Nahavand University, Nahavand, 6593139565, Iran.
Department of Biotechnology, College of Agriculture, Bu-Ali Sina University, Hamadan, 65167, Iran.
Anal Bioanal Chem. 2018 Jun;410(16):3683-3691. doi: 10.1007/s00216-018-1005-y. Epub 2018 Apr 7.
Sensitive detection of biomarkers will mean accurate and early diagnosis of diseases. A tissue plasminogen activator (tPA) has a crucial role in many cardiovascular diseases and it is related to many processes such as angiogenesis in cancer cells. Therefore, sensitive determination of tPA is important in diagnosis and clinical research. tPA monoclonal antibody was covalently attached onto single-wall carbon nanotubes (SWCNTs) using diimide-activated imidation coupling. Functionalized SWCNTs were immobilized onto a glassy carbon electrode and the modification process was investigated by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE), SEM, cyclic voltammetry (CV), and electrochemical impedance spectroscopy (EIS). Cyclic voltammograms (CVs) in a scan rate of 100 mVs was studied and comparisons were made between the modified glassy carbon electrodes (immobilized with antibodies) as a working electrode before and after the formation of tPA-antibody complex. Results of the SDS-PAGE demonstrated that the antibody was covalently and site directly attached to the SWCNTs. The fabricated biosensor provided a good linear response range from 0.1 to 1.0 ng mL with a low detection limit of 0.026 ng mL. The immunosensor showed selectivity, reproducibility, good sensitivity, and acceptable stability. Satisfactory results were observed for early and sensitive determination of tPA in human serum samples. For the first time, such specific biosensor is currently being fabricated for tPA in our laboratories and successfully could determine tPA in myocardial infraction and breast cancer patients. Graphical abstract Fabricated biosensor for determination of tPA.
生物标志物的灵敏检测意味着对疾病的准确和早期诊断。组织型纤溶酶原激活物（tPA）在许多心血管疾病中起着至关重要的作用，它与许多过程有关，如癌细胞中的血管生成。因此，tPA 的灵敏测定在诊断和临床研究中很重要。使用二酰亚胺激活的酰亚胺偶联将组织型纤溶酶原激活物单克隆抗体共价连接到单壁碳纳米管（SWCNTs）上。将功能化的 SWCNTs 固定在玻碳电极上，并通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（SDS-PAGE）、SEM、循环伏安法（CV）和电化学阻抗谱（EIS）研究修饰过程。在 100 mV/s 的扫描速率下研究了循环伏安图（CV），并比较了在形成 tPA-抗体复合物之前和之后作为工作电极的修饰玻碳电极。SDS-PAGE 的结果表明，抗体被共价且直接连接到 SWCNTs 上。所制备的生物传感器提供了从 0.1 到 1.0 ng mL 的良好线性响应范围，检测限低至 0.026 ng mL。该免疫传感器具有选择性、重现性、良好的灵敏度和可接受的稳定性。在人血清样品中对 tPA 的早期和灵敏测定观察到了令人满意的结果。这是我们实验室首次为 tPA 制造这种特定的生物传感器，并成功地确定了心肌梗死和乳腺癌患者的 tPA。
