Patel Rhea, Vinchurkar Madhuri, Mohin Shaikh Aatha, Patkar Rajul, Adami Andrea, Giacomozzi Flavio, Ramesh Raman, Pramanick Bidhan, Lorenzelli Leandro, Shojaei Baghini Maryam
Center for Research in Nanotechnology and Science, Indian Institute of Technology Bombay, Mumbai 400076, India.
Department of Electrical Engineering, Indian Institute of Technology Bombay, Mumbai 400076, India.
Bioelectrochemistry. 2023 Apr;150:108370. doi: 10.1016/j.bioelechem.2023.108370. Epub 2023 Jan 7.
Herein, we report for the first time the development of a label-free, non-faradaic, and highly sensitive DNA-based impedimetric sensor using micro-sized gold interdigitated electrodes (IDE) to detect a soil-borne agricultural pathogen Ralstonia solanacearum. A universal 30 oligomer single-stranded DNA (ssDNA) probe lpxC4 having specificity towards R. solanacearum is successfully immobilized on the surface of IDE along with mercaptohexanol. The electrochemical stability of the developed sensor surface is determined using open circuit potential measurements. The DNA probe immobilization protocol is validated using the changes configured on the surface of IDE by contact angle and ATR-FTIR analysis. The DNA target hybridization is detected using non-faradaic electrochemical impedance spectroscopy measurement with an ultra-low sample volume of 10 µL. The non-faradaic approach is verified by studying redox behavior using cyclic voltammetry. We investigate the hybridization of the surface-immobilized label-free probe with the complementary DNA targets obtained from infected eggplant saplings and cross-reactive studies with mismatched DNA strands. Our impedimetric sensor can detect target concentrations as low as 0.1 ng/µL. This standardization and detection of DNA hybridization serves as part I of the work and paves the way for further study in the detection of pathogenic field samples.
在此，我们首次报道了一种基于微尺寸金叉指电极（IDE）的无标记、非法拉第且高灵敏度的基于DNA的阻抗传感器的研制，用于检测一种土传农业病原菌青枯雷尔氏菌。一种对青枯雷尔氏菌具有特异性的通用30聚体单链DNA（ssDNA）探针lpxC4与巯基己醇一起成功固定在IDE表面。通过开路电位测量来确定所研制传感器表面的电化学稳定性。利用接触角和衰减全反射傅里叶变换红外光谱（ATR-FTIR）分析对IDE表面配置的变化进行验证，从而对DNA探针固定方案进行验证。使用超低样品体积为10 μL的非法拉第电化学阻抗谱测量来检测DNA靶标杂交。通过循环伏安法研究氧化还原行为来验证非法拉第方法。我们研究了表面固定的无标记探针与从感染茄子幼苗中获得的互补DNA靶标的杂交以及与错配DNA链的交叉反应研究。我们的阻抗传感器能够检测低至0.1 ng/μL的靶标浓度。DNA杂交的这种标准化和检测作为该工作的第一部分，为进一步研究病原菌现场样品的检测铺平了道路。
