Development and Functionalization of Electrolyte-Gated Graphene Field-Effect Transistor for Biomarker Detection
Summary
The present protocol demonstrates the development of electrolyte-gated graphene field-effect transistor (EGGFET) biosensor and its application in biomarker immunoglobulin G (IgG) detection.
Abstract
In the current study, graphene and its derivatives have been investigated and used for many applications, including electronics, sensing, energy storage, and photocatalysis. Synthesis and fabrication of high quality, good uniformity, and low defects graphene are critical for high-performance and highly sensitive devices. Among many synthesis methods, chemical vapor deposition (CVD), considered a leading approach to manufacture graphene, can control the number of graphene layers and yield high-quality graphene. CVD graphene needs to be transferred from the metal substrates on which it is grown onto insulating substrates for practical applications. However, separation and transferring of graphene onto new substrates are challenging for a uniform layer without damaging or affecting graphene's structures and properties. Additionally, electrolyte-gated graphene field-effect transistor (EGGFET) has been demonstrated for its wide applications in various biomolecular detections because of its high sensitivity and standard device configuration. In this article, poly (methyl methacrylate) (PMMA)-assisted graphene transferring approach, fabrication of graphene field-effect transistor (GFET), and biomarker immunoglobulin G (IgG) detection are demonstrated. Raman spectroscopy and atomic force microscopy were applied to characterize the transferred graphene. The method is shown to be a practical approach for transferring clean and residue-free graphene while preserving the underlying graphene lattice onto an insulating substrate for electronics or biosensing applications.
Introduction
Graphene and its derivatives have been investigated and used for many applications, including electronics1,2, sensing3,4,5, energy storage6,7, and photocatalysis1,6,8. Synthesis and fabrication of high quality, good uniformity, and low defects graphene are critical for high-performance and highly sensitive devices. Since the development of Chemical vapor deposition (CVD) in 2009, it has shown colossal promise and set its place as an essential member of the graphene family9,10,11,12,13. It is grown on a metal substrate and, later for practical uses, is transferred onto insulating substrates14. Several transferring methods have been used to transfer CVD graphene recently. The poly (methyl methacrylate) (PMMA) assisted method is the most used among the different techniques. This method is particularly well-suited for industrial usage because of its large-scale capability, lower cost, and high quality of the transferred graphene14,15. The critical aspect of this method is getting rid of the PMMA residue for CVD graphene's applications because the residues can cause declination of the electronic properties of graphene14,15,16, cause an effect on biosensors' sensitivity and performance17,18, and create significant device-to-device variations19.
Nanomaterials-based biosensors have been significantly investigated over the past decades, including silicon nanowire (SiNW), carbon nanotube (CNT), and graphene20. Because of its single-atom-layer structure and distinctive properties, graphene demonstrates superior electronic characteristics, good biocompatibility, and facile functionalization, making it an attractive material for developing biosensors14,21,22,23. Due to field-effect transistors (FET) characteristics such as high sensitivity, standard configuration, and cost-effective mass producibility21,24, FET is more preferred in portable and point-of-care implementations than other electronics-based biosensing devices. The electrolyte-gated graphene field-effect transistor (EGGFET) biosensors are examples of the stated FETs21,24. EGGFET can detect various targeting analytes such as nucleic acids25, proteins24,26, metabolites27, and other biologically relevant analytes28. The technique mentioned here ensures the implementation of CVD graphene in a label-free biosensing nanoelectronics device which offers higher sensitivity and accurate time detection over other biosensing devices29.
In this work, an overall process for developing an EGGFET biosensor and functionalizing it for biomarker detection, including transferring CVD graphene onto an insulating substrate, Raman, and AFM characterizations of the transferred graphene, are demonstrated. Furthermore, fabrication of EGGFET and integration with a polydimethylsiloxane (PDMS) sample delivery well, bioreceptor functionalization, and successful detection of human immunoglobulin G (IgG) from serum by spike-and-recovery experiments are also discussed here.
Protocol
Representative Results
Discussion
The purchased CVD graphene on copper film needs to be trimmed to the right size for the following fabrication steps. Cutting of the films can cause wrinkling, which needs to be prevented. The parameters provided in the fabrication step can be referred to for plasma etching of graphene, and these numbers could be varied when using different instruments. The etched sample must be closely monitored and inspected to ensure complete graphene etching. Multiple pre-cleaning methods can be applied to clean the substrates, such a…
Disclosures
The authors have nothing to disclose.
Acknowledgements
The experiments were conducted at West Virginia University. We acknowledge the Shared Research Facilities at West Virginia University for device fabrication and material characterization. This work was supported by the US National Science Foundation under Grant No. NSF1916894.
Materials
| 1-pyreneutyric acid N- hydroxysuccinimide ester | Sigma Aldrich | 457078-1G | functionalization |
| Asylum MFP-3D Atomic Force Microscope | Oxford Instruments | graphene characterization | |
| AZ 300 MIF | MicroChemicals | AZ 300 MIF | photoresist developer |
| AZ 300 MIF | MicroChemicals | AZ 300 MIF | photoresist |
| Bovine Serum Albumin | Sigma Aldrich | 810014 | blocking |
| Branson 1210 Sonicator | SONITEK | sample cleaning | |
| Copper Etchant | Sigma Aldrich | 667528-500ML | removing copper film to release graphene |
| Dimethyl Sulfoxide (DMSO) | VWR | 97063-136 | functionalization |
| Disposable Biopsy Punches, Integra Miltex | VWR | 21909-144 | create well in PDMS |
| Gold etchant | Gold Etch, TFA, Transene | 658148 | enchant |
| Graphene | Graphene supermarket | 2" x 2" sheet | biosensing element of the device |
| IgG aptamer | Base Pair Biotechnologies | customized | bioreceptor |
| Keithley 4200A-SCS Parameter Analyzer | Tektronix | measurement and detection | |
| KMG CR-6 | KMG chemicals | 64216 | Chromium etchant |
| Kurt J. Lesker E-beam Evaporator | Kurt J. Lesker | metal deposition | |
| Laurell Technologies 400 Spinners | Laurell Technologies | WS-400BZ-6NPP/LITE | thin film coating |
| March PX-250 Plasma Asher | March Instruments | sample cleaning | |
| Nickel etchant | Nickel Etchant, TFB, Transene | 600016000 | etchant |
| OAI Flood Exposure | OAI | photolithography | |
| Phosphate Buffered Saline (PBS) | Sigma Aldrich | 806552-500ML | buffer |
| PMMA 495K A4 | MicroChemicals | PMMA 495K A4 | Photoresist for assisting graphene transferring |
| Polydimethylsiloxane (PDMS) | Sigma Aldrich | Sylgard 184 | sample delivery well |
| Renishaw InVia Raman Microscope | Renishaw | graphene characterization | |
| Sodium Hydroxide (NaOH) | Sigma Aldrich | 221465-25G | functionalization |
| Suss Microtech MA6 Mask Aligner | Suss MicroTec | photolithography | |
| Thermo Scientific Cimarec Hotplate | Thermo Scientific | SP131635 | sample and device Baking |
References
- Saini, D. Synthesis and functionalization of graphene and application in electrochemical biosensing. Nanotechnology Reviews. 5 (4), 393-416 (2016).
- Emtsev, K. V., Bostwick, A., Horn, K., et al. Towards wafer-size graphene layers by atmospheric pressure graphitization of silicon carbide. Nature Materials. 8 (3), 203-207 (2009).
- Wang, Y., et al. Electrochemical delamination of CVD-grown graphene film: Toward the recyclable use of copper catalyst. ACS Nano. 5 (12), 9927-9933 (2011).
- Carvalho Fernandes, D. C., Lynch, D., Berry, V. 3D-printed graphene/polymer structures for electron-tunneling based devices. Scientific Reports. 10 (1), 1-8 (2020).
- Gao, L., et al. Repeated growth and bubbling transfer of graphene with millimetre-size single-crystal grains using platinum. Nature Communications. 3, 699 (2012).
- Singh, J., Rathi, A., Rawat, M., Gupta, M. Graphene: From synthesis to engineering to biosensor applications. Frontiers of Materials Science. 12 (1), 1-20 (2018).
- Randviir, E. P., Brownson, D. A. C., Banks, C. E. A decade of graphene research: Production, applications and outlook. Materials Today. 17 (9), 426-432 (2014).
- Suvarnaphaet, P., Pechprasarn, S. Graphene-based materials for biosensors: A review. Sensors (Switzerland). 17 (10), 2161 (2017).
- Li, X., Cai, W., An, J., et al. Large-area synthesis of high-quality and uniform graphene films on copper foils. Science. 324 (5932), 1312-1314 (2009).
- Yu, Q., Lian, J., Siriponglert, S., Li, H., Chen, Y. P., Pei, S. S. Graphene segregated on Ni surfaces and transferred to insulators. Applied Physics Letters. 93 (11), 113103 (2008).
- Xu, S. C., et al. Direct synthesis of graphene on SiO2 substrates by chemical vapor deposition. CrystEngComm. 15 (10), 1840-1844 (2013).
- Zhang, C., et al. Facile synthesis of graphene on dielectric surfaces using a two-temperature reactor CVD system. Nanotechnology. 24 (39), 395603 (2013).
- Zhang, C., et al. Direct formation of graphene-carbon nanotubes hybrid on SiO2 substrate via chemical vapor deposition. Science of Advanced Materials. 6 (2), 399-404 (2014).
- Sun, J., Finklea, H. O., Liu, Y. Characterization and electrolytic cleaning of poly(methyl methacrylate) residues on transferred chemical vapor deposited graphene. Nanotechnology. 28 (12), 125703 (2017).
- Lin, Y. C., Lu, C. C., Yeh, C. H., Jin, C., Suenaga, K., Chiu, P. W. Graphene annealing: How clean can it be. Nano Letters. 12 (1), 414-419 (2012).
- Pirkle, A., et al. The effect of chemical residues on the physical and electrical properties of chemical vapor deposited graphene transferred to SiO2. Applied Physics Letters. 99 (12), 122108 (2011).
- Chen, T. Y., et al. Label-free detection of DNA hybridization using transistors based on CVD grown graphene. Biosensors and Bioelectronics. 41 (1), 103-109 (2013).
- Xu, S., et al. Direct growth of graphene on quartz substrates for label-free detection of adenosine triphosphate. Nanotechnology. 25 (16), 165702 (2014).
- Dan, Y., Lu, Y., Kybert, N. J., Luo, Z., Johnson, A. T. C. Intrinsic response of graphene vapor sensors. Nano Letters. 9 (4), 1472-1475 (2009).
- Zhang, A., Lieber, C. M. -. Nano-Bioelectronics. Chemical Reviews. 116 (1), 215-257 (2015).
- Forsyth, R., Devadoss, A., Guy, O. J. Graphene Field effect transistors for biomedical applications: Current status and future prospects. Diagnostics (Basel). 7 (3), 45 (2017).
- Dankerl, M., et al. Graphene solution-gated field-effect transistor array for sensing applications. Advanced Functional Materials. 20 (18), 3117-3124 (2010).
- He, Q., Wu, S., Yin, Z., Zhang, H. Graphene -based electronic sensors. Chemical Science. 3 (6), 1764-1772 (2012).
- Sun, J., Liu, Y. Matrix effect study and immunoassay detection using electrolyte-gated graphene biosensor. Micromachines. 9 (4), 142 (2018).
- Mohanty, N., Berry, V. Graphene-based single-bacterium resolution biodevice and DNA transistor: Interfacing graphene derivatives with nanoscale and microscale biocomponents. Nano Letters. 8 (12), 4469-4476 (2008).
- Ohno, Y., Maehashi, K., Yamashiro, Y., Matsumoto, K. Electrolyte-gated graphene field-effect transistors for detecting pH and protein adsorption. Nano Letters. 9 (9), 3318-3322 (2009).
- Huang, Y., Dong, X., Shi, Y., Li, C. M., Li, L. J., Chen, P. Nanoelectronic biosensors based on CVD grown graphene. Nanoscale. 2 (8), 1485-1488 (2010).
- Jiang, S., et al. Real-time electrical detection of nitric oxide in biological systems with sub-nanomolar sensitivity. Nature Communications. 4 (1), 1-7 (2013).
- Bai, Y., Xu, T., Zhang, X. Graphene-based biosensors for detection of biomarkers. Micromachines. 11 (1), 60 (2020).
- Madou, M. J. . Fundamentals of Microfabrication The Science of Miniaturization. 2nd ed. , (2002).
- Xia, Y., Whitesides, G. M. Soft lithography. Annual Review of Material Sciences. 28 (1), 153-184 (2003).
- Wang, Y. Y., et al. Raman studies of monolayer graphene: The substrate effect. Journal of Physical Chemistry C. 112 (29), 10637-10640 (2008).
- Betancur, V., Sun, J., Wu, N., Liu, Y. Integrated lateral flow device for flow control with blood separation and biosensing. Micromachines. 8 (12), 367 (2017).
- Butt, A. . Physics and Chemistry of Interfaces. 3rd ed. , (2003).
- Sitko, R., Zawisza, B., Malicka, E. Graphene as a new sorbent in analytical chemistry. TrAC Trends in Analytical Chemistry. 51, 33-43 (2013).
- Bai, L., et al. Graphene for energy storage and conversion: Synthesis and Interdisciplinary applications. Electrochemical Energy Reviews. 3 (2), 395-430 (2019).
- Boretti, A., Al-Zubaidy, S., Vaclavikova, M., Al-Abri, M., Castelletto, S., Mikhalovsky, S. Outlook for graphene-based desalination membranes. npj Clean Water. 1 (1), 1-11 (2018).
- Pumera, M. Graphene in biosensing. Materials Today. 14 (7-8), 308-315 (2011).
- Sun, J., Liu, Y. Unique constant phase element behavior of the electrolyte-graphene interface. Nanomaterials. 9 (7), 923 (2019).
- Sun, J., Camilli, L., Caridad, J. M., Santos, J. E., Liu, Y. Spontaneous adsorption of ions on graphene at the electrolyte-graphene interface. Applied Physics Letters. 117 (20), 203102 (2020).
