方法文章

High-Performance Graphene-Modified Sensing Chip for SARS-CoV-2 Detection

DOI：

10.3791/64730

⸱

2023年5月5日

 ,  ,  ,  , 
1RFIC Bio Centre, Kwangwoon University, 2Department of Electronics Engineering, Kwangwoon University, 3NDAC Centre, Kwangwoon University, 4Stanford Cardiovascular Institute, Stanford University School of Medicine, 5Department of Medicine, Stanford University School of Medicine, 6Department of Biomedical Engineering, Hebei University of Technology, 7NanoBioTech Laboratory, Department of Environmental Engineering, Florida Polytechnic University, 8School of Engineering, University of Petroleum and Energy Studies (UPES)

本文内容

摘要

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The present protocol describes the fabrication of low-cost biosensing prototypes based on useful nanosystems for accurately detecting viral proteins (at the Fg level). Such a tiny sensor platform allows for point-of-care applications that can be integrated with the Internet of Medical Things (IoMT) to meet telemedicine objectives.

摘要

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This sensing prototype model involves the development of a reusable, twofold graphene oxide (GrO)-glazed double inter-digitated capacitive (DIDC) detecting chip for detecting severe acute respiratory syndrome coronavirus 2 virus (SARS-CoV-2) specifically and rapidly. The fabricated DIDC comprises a Ti/Pt-containing glass substrate glazed with graphene oxide (GrO), which is further chemically modified with EDC-NHS to immobilize antibodies (Abs) hostile to SARS-CoV-2 based on the spike (S1) protein of the virus. The results of insightful investigations showed that GrO gave an ideal engineered surface for Ab immobilization and enhanced the capacitance to allow higher sensitivity and low sensing limits. These tunable elements helped accomplish a wide sensing range (1.0 mg/mL to 1.0 fg/mL), a minimum sensing limit of 1 fg/mL, high responsiveness and good linearity of 18.56 nF/g, and a fast reaction time of 3 s. Besides, in terms of developing financially viable point-of-care (POC) testing frameworks, the reusability of the GrO-DIDC biochip in this study is good. Significantly, the biochip is specific against blood-borne antigens and is stable for up to 10 days at 5 °C. Due to its compactness, this scaled-down biosensor has the potential for POC diagnostics of COVID-19 infection. This system can also detect other severe viral diseases, although an approval step utilizing other virus examples is under development.

引言

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A viral pandemic caused by a new beta coronavirus1 (i.e., 2019-nCoV), which was later named as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)2 (hereafter predominantly referred to as the virus), involving a pneumonic cluster and severe acute respiratory distress, emerged in the Wuhan city, China, at the end of 20193. Owing to its fast worldwide human-to-human transmission, high infection rate, high mortality rate, and serious life-threatening adverse effects4, during the pandemic, virology research5 evolved quickly to identify....

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方案

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1. Cleaning of the DIDC sensing chip

  1. At the beginning of the experiment, clean the DIDC chip surface26 with piranha solution (H2SO4:H2O2in a 3:1 ratio), and place it on the hot plate at 80 °C for 15 min. Next, rinse the sensor surface with distilled water drop by drop using a pipette to remove the cleaning reagents completely. To ensure the complete removal of the reagent, rinse the surface with four to five drops of ethyl alcohol.
    NOTE: The DIDC chip was fabricated following a previously published report26.
  2. Then, dry the ....

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结果

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Here, a protocol is presented for sensing the S1 protein of the SARS-CoV-2 virus using a graphene oxide-glazed double inter-digitated capacitive (DIDC) sensing chip. Figure 1 shows a schematic representation (fabrication with the circuit layout) of the extremely sensitive and recyclable graphene oxide-modified double interdigitated capacitive (DIDC) sensing chip. The detailed stepwise fabrication process is shown in Figure 2. Figure 3

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讨论

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For fashioning a productive DIDC chip-based biosensor, the charge distribution, conductivity, and dielectric constant of the DIDC are extremely important. Significantly, the improvements in these detection boundaries relate to the capacitive reactance of the DIDC18,26,27. In this study, a capacitance immunosensor was fabricated that is hostile to the virus Abs and functionalized by EDC-NHS coupling on the graphene oxide-DIDC-bas.......

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披露

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The authors have nothing to disclose.

致谢

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This work was upheld to some extent by the Basic Science Research Program through the National Research Foundation of Korea (NRF) sponsored by the Ministry of Education under Grant 2018R1D1A1A09083353 and Grant 2018R1A6A1A03025242, somewhat by the GCS Group Association Ltd., and by the Korea Ministry of Environment (MOE) Graduate School invested huge energy in Integrated Pollution Prevention and Control Project and a Research Grant of Kwangwoon University in 2022.

E.M. would like to acknowledge the support from the National Institute of Biomedical Imaging and Bioengineering (5T32EB009035).

....

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材料

本文使用的材料清单
姓名公司目录编号评论
淀粉样蛋白和 β;1-42 蛋白默克 （Sigma-Aldrich）107761-42-2
抗 SARS-CoV-2 刺突 （S1） 单克隆 IgG 抗体 义翘神州40150-R007
EDC [N-（3-二甲氨基丙基）-N′-乙基碳二亚胺盐酸盐]赛默飞世尔科技A35391
乙醇（C2H5OH）Sigma-Aldrich
过氧化氢（H2O2）
酰
NHS（NHydroxysuccinimide，98+%;C4H5NO3）Thermo Fisher ScientificA39269
PBS
前列腺特异性抗原 Sigma-AldrichP3338-25UG
SARS-CoV-2 刺突 S1-His 重组蛋白义翘神州40591-V08H
单层氧化石墨烯石墨烯超市
旋涂机高精度旋涂机（旋涂系统）ACE-200 
硫酸 （H2SO4）
聚亚胺带

参考文献

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  1. Boldog, P. Risk assessment of novel coronavirus COVID-19 outbreaks outside China. Journal of Clinical Medicine. 9 (2), 571-583 (2020).
  2. Seo, G., et al. Rapid detection of COVID-19 cau....

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