Applied Science and Convergence Technology 2022; 31(4): 89-92
Published online July 30, 2022
Copyright © The Korean Vacuum Society.
aInfectious Disease Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon 34141, Republic of Korea
bDepartment of Biotechnology, Korea University of Science and Technology (UST), Daejeon 34141, Republic of Korea
Serotonin is a stress biomarker and is one of the major neurotransmitters, and its concentration in the body is related to psychological functions such as mental illness. In particular, this biomarker is used to indicate depression caused by the increased stress of modern society. Therefore, detection and monitoring technologies are important for tracing concentration changes. In this study, we developed a serotonin antibodyconjugated graphene micropatterned field-effect transistor (SAb-GMFET)-based portable device for on-site or self-diagnosis of serotonin. The SAb-GMFET consisted of a graphene micropatterned channel, SAb for selective detection, and electrodes to supply the voltage. The SAb was immobilized by forming an amide bond with a diamine linker. SAb-GFMET showed high performance of limit of detection of 10 pM within 10 s and exhibited excellent selective detection among the interfering materials. For on-site and self-diagnosis applications, a portable device was developed with a sim chip, and the SAb-GMFET was connected to a printed circuit board using wire bonding. The portable SAb-GMFET showed a similar performance to that of the SAb-GMFET. Therefore, this portable platform can be utilized for point-of-care tests.
Keywords: Serotonin, Portable biosensor, Graphene, Field-effect transistor, Neurotransmitters
Serotonin is one of the monoamine neurotransmitters in the central nervous system, and its concentration is important in the body owing to its responses to physiological and psychological functions, such as sleep, Alzheimer’s disease, appetite, pain, sexuality, desire, and mental state [1,2]. Generally, the concentration of serotonin in the human body is approximately 295 to 687 ng/mL in human urine, and irregular concentrations can occur with various disorders [3,4]. In particular, an increasing number of people have suffered depression owing to numerous stresses from their personal lives, society, and the COVID-19 pandemic. Therefore, rapid detection methodologies are required for monitoring changes in serotonin concentration.
Although many technologies, including cyclic voltammetry, radioimmunoassays, chemiluminescence, enzyme-linked immunosorbent assay, field-effect transistors, fluorescence, and the polymerase chain reaction, have been developed for serotonin analyses, these methods have limitations; they can be time-consuming or exhibit a low detection range, low sensitivity, and low selectivity [1,5–10]. Although various electronic sensors were recently developed for serotonin detection, these sensor platforms show deficiencies related to rapid detection, real-time monitoring, and on-site application systems. To overcome these limitations, technology is required to provide high performance, quick testing, a broad detection range, high-sensitivity and selective detection, and this remains a challenge.
In this study, we first demonstrate a real-time monitoring system using a portable serotonin antibody (SAb)-graphene micropatterned field-effect transistor (GMFET) device for on-site monitoring and serotonin detection. Graphene micropatterned (GM) was used with SAb as a bioprobe for the channel of the FET, and the selective serotonin detector bis(2-aminoethylene)perylene-3,4,9,10-tetracarboxyldiimide (PDA) was utilized to chemically immobilize the SAb onto the graphene. In particular, PDA was synthesized to include two amine groups at the terminus for increased conjugation with SAb . The SAb-GMFET showed high sensitivity and selectivity, a limit of detection (LOD) of 10 pM and a wide range for 10 pM to 100 nM with a 10 s response time. Moreover, the
Copper foil and glutaraldehyde (GA) were purchased from Thermo Fisher and Sigma Aldrich Co., Ltd. Samples of 950 PMMA A4 4 % and serotonin antibody (ab66047) were purchased from Micro Chem Co., Ltd. and Abcam. Selected area electron diffraction (SAED) images were obtained using high-resolution transmission electron microscopy (Jeol, JEM-ARM200F), and Raman spectra were obtained with a LabRAM HR Evolution Visible_NIR system from HORIBA using a 633 nm laser.
Monolayer graphene (MLG) was synthesized using a chemical vapor deposition (CVD) method based on our previous reports [12–15]. Graphene film was formed on the copper foil substrate and was synthesized from the gas sources CH4, H2, and Ar at 1,000 °C. Then, the MLG surface was coated with PMMA for protection. The copper film was removed using copper etchant, and the MLG was transferred onto a silicon dioxide wafer using the wet-transfer method. Finally, the PMMA was removed by acetone vapor and washed with distilled water.
The electrodes for the SAb-GMFET were fabricated by a microelectromechanical system (MEMS) process [16,17]. AZ GNX-601 positive photoresist was utilized with GM in the photolithography process, and a reactive-ion etching system was used for graphene etching. A DNR L300-40 negative photoresist was used to form the electrode (Cr/Au) and passivation layer (SiO2). Ten micromolar PDA linker compound was used for conjugation on the GM channel for 30 min, and 2 % GA coupling agent was dripped onto the PDA/GM for 4 h. SAb with a concentration of 2 µg/mL was reacted with the modified GM channel for 4 h at 4 °C.
All electrical measurements were performed using a probe station (MS-tech, Model 4000) and a Keithley 2612 A source meter. The normalized sensitivity was calculated with the following equation:
MLG has various properties, such as a zero bandgap, high carrier mobility, high conductivity, ambipolar properties, and rapid electron transport [14,18–20]. MLG has been utilized in various applications and has been applied in preparing bio/chemosensors owing to its excellent electrical properties [21,22]. To confirm the preparation of MLG, the graphene sample was analyzed using high-resolution transmission electron microscopy (HR-TEM), and the hexagonal lattice pattern was observed in a SAED image [Fig. 1(a)]. The MLG surface was modified by PDA for immobilization of the SAb, and the PDA interfacial chemical was stacked on graphene via π interactions between the
Current-voltage (I-V) curves were measured over the range −2 V to 2 V, and the graphs showed gradually decreasing slopes that depended on the surface modifications of the MLG [Fig. 2(a)]. However, the current was slightly decreased by π-π stacking due to PDA introduction (physical attachment) and conjugation of the SAb (chemical attachment), which resulted in obvious ohmic properties and followed Ohm’s law. An illustrated scheme is presented for the liquid-ion gated FET system with the SAb conjugated-GMFET, which consisted of source (S), drain (D), and gate (G) electrodes [Fig. 2(b)]. In addition, PBS solution was added into the chamber as a dielectric material, which was formed by the field effect of the voltage supplied from the gate electrode. Following our previous studies, the conductance of the GMFET showed ambipolar properties. The liquid-ion gated FET system was suitably verified of the operation in the
To confirm the serotonin detection performance of the SAb-GMFET, an electrode was fabricated by using the MEMS process. The liquidion gated SAb-GMFET was tested during exposure in real time, and the sensitivity of the detection performance was calculated with the normalization based following equation:
The real-time responses of the SAb-GMFET to various concentrations of serotonin are displayed in Fig. 3(a). Before monitoring the current change, a buffer solution without serotonin was injected to confirm the buffer effect, and there was no electrical change. On the other hand, changes in the current were immediately observed after exposure to serotonin in the concentration range 10 pM to 100 nM, and the response time was less than 10 s. The LOD of the SAb-GMFET was 10 pM, which was 30 times higher than those seen in other studies using SAb (Table I). The concentration curve was obtained from the normalized sensitivity shown in Fig. 3(a). A linear response was displayed in the concentration range 10 pM to 10 nM, and the current level was saturated at concentrations over 100 nM [Fig. 3(b)]. To investigate the binding affinity, the
Although many serotonin detection technologies have been developed, these analysis methodologies exhibit limitations such as low sensitivity, low selectivity, and long measurement times. The SAb-GMFET platform was utilized in a portable device for PoCT health care applications [Fig. 4(a)]. The portable platform consisted of a hardware platform based on a PCB, such as a microcontroller unit (MCU), power supply, communication module for outgoing data, and chip socket for docking of the chip integrated with the SAb-GMFET electrode. In addition, the sim chip with the integrated SAb-GMFET was connected by wire bonding between the PCB and electrode to supply the voltage. The performance of the portable SAb-GMFET was evaluated by injecting the LOD concentration of serotonin [Fig. 4(b)]. The observed sensitivity was similar to that seen for the SAb-GMFET, as shown in Fig. 3(a), which indicates the possibility of on-site utilization. In addition, a portable SAb-GMFET was used to carry out real-time monitoring of the diluted serotonin in artificial urine for possible selfdiagnosis [Fig. 4(c)]. The results were normalized and compared with those in Fig. 4(b), and they showed similar results for the same concentrations.
In summary, we have demonstrated a sensitive and selective SAb-GMFET platform and developed a portable device. The SAb-GMFET achieved rapid detection and real-time monitoring of serotonin and comprised SAb for selective detection and a liquid-ion gated FET as a highly sensitive monitoring system. The SAb-GMFET exhibited excellent sensitivity with an LOD of 10 pM. In addition, the device demonstrated selective detection of serotonin at concentrations 100 times lower than those of various interfering materials, such as neurotransmitters and hormones. Moreover, a portable SAb-GMFET device was constructed by integration of the SAb-GMFET with a sim and connection using wire bonding. The portable device showed excellent detection of serotonin in artificial urine. Therefore, this portable SAb- GMFET platform can be utilized for PoCT and self-diagnosis.
This research was supported by Research Program to Solve Urgent Safety Issues of the National Research Foundation of Korea (NRF) funded by the Korean government (Ministry of Science and ICT (MSIT)) (NRF-2020M3E9A1111636); Korea Institute of Planning and Evaluation for Technology in Food, Agriculture and Forestry (IPET) and Korea Smart Farm R&D Foundation (KosFarm) through Smart Farm Innovation Technology Development Program, funded by Ministry of Agriculture, Food and Rural Affairs (MAFRA) and Ministry of Science and ICT (MSIT), Rural Development Administration (RDA) (421020- 03); the National Research Council of Science & Technology (NST) grant by the Korea government (MSIT) (No. CAP22011-000); the National R&D Program of National Research Foundation of Korea (NRF) funded by Ministry of Science and ICT (2021M3H4A4079276); the Korea Research Institute of Bioscience and Biotechnology (KRIBB) Research Initiative Program (1711170578).