EC methods are powerful tools for creating sensitive and selective sensing platforms owing to several properties such as rapid detection time, convenience, and stability. To fabricate EC sensors to detect DA release in living cells, key factors need to be satisfied with the electrode surface materials. First, excellent electrical properties, including electrical conductivity and electron mobility, accelerate the redox reaction as a critical mechanism in EC sensing. Second, biocompatibility does not inhibit the adhesion, function, and growth of neuronal cells that adhere to the surface. In this section, we categorise the materials into three different areas: (i) Carbon-based materials, (ii) hybrid gold nanoparticles, and (iii) bare gold structures with different geometries.
Carbon-based materials for the development of electrochemical dopamine sensors
Carbon nanomaterials, a class of materials mainly composed of carbon (i.e., the graphene family, CNTs, and carbon nanofibers (CNFs)), have been utilised in various fields due to their unique properties such as thermal stability, chemical resistance, and excellent mechanical properties [66,67,68,69]. Generally, most carbon-based materials have a specific chemical structure, where carbon atoms combine via SP2 hybridisation, which causes delocalisation of electrons in their structure [70]. Owing to their structural features, they have excellent electrical properties, including electrical conductivity and electron mobility. In addition, carbon-based nanomaterials with enhanced electrical ability and additional capabilities can be obtained through chemical or physical modification [71]. Given that biosensing based on EC detection techniques (i.e., cyclic voltammetry, amperometry, and differential pulse voltammetry) requires a platform with excellent electrical properties, carbon-based nanomaterials have tremendous potential for sensing target biomolecules with their redox activities.
Graphene family-based dopamine (DA) sensors
To date, the graphene family has been widely used for the electrical detection of DA with the modification of other chemicals such as gold nanoparticles. In addition, graphene is controllable in terms of shape, size, and structure. Recently, Zan et al. reported that a reduced graphene oxide-based EC sensing platform is highly effective in detecting DA released from pheochromocytoma cells [51]. In this study, a reduced graphene oxide composite was fabricated with a flexible ultra-thin structure. Moreover, the graphene composite was modified with dendritic platinum nanoparticles, which increased the sensitivity of sensing DA in living cells. To fabricate the sensing platform, ultra-thin reduced graphene oxide was prepared by sequential mould casting, a method suitable for controlling the size and thickness of graphene and chemical reduction. Subsequently, the reduced graphene oxide composite was decorated with platinum nanoparticles synthesised by thermal synthesis through the dip-coating method. Based on the results of EC detection, such as differential pulse voltammetry (DPV) and cyclic voltammetry (CV), it was confirmed that this sensing platform could recognise EC signals from DA in the presence of interfering molecules (i.e., ascorbic acid and uric acid). Furthermore, it was demonstrated that the sensing platform is adequate for cell cultivation and the detection of DA secreted from living cells. In 2017, Emran et al. developed a novel carbon material-based amperometric sensing platform for monitoring DA from living cells (Fig. 2a) [52].
Interestingly, the carbon materials used in this study have unique structures and properties, specifically, three-dimensional N-doped mesoporous nanosheets with excellent biocompatibility. Owing to their structural and chemical features, the sensing platform has several advantages: (i) high surface area, (ii) excellent biocompatibility, and (iii) enhanced electrocatalytic features for highly sensitive detection of DA released from living cells, which leads to improved detection capability (a linear range of < 0.5 mM and a limit of detection (LOD) of 10 nM), as compared with that of conventional sensing electrodes. Moreover, the amperometric responses from chemical DA were selectively recognised in the presence of several interfering molecules (Fig. 2b). Above all, it was observed that DA from living cells was detectable under K + stimulation through amperometric responses (Fig. 2c). In 2019, Asif et al. introduced highly sensitive and selective DA sensors composed of hetero-stacked layers, including reduced graphene oxide (rGO) and layered double hydroxides (LDHs) [53]. Specifically, the LDHs improved the catalytic capability of the composite. Additionally, the rGO layer was hybridised with an extremely thin LDH layer. It was fabricated on a single molecular scale to improve both the surface area of the composite and the efficacy of electrocatalytic activity. According to the CV measurements, it was observed that the electrochemical peaks of ascorbic acid (AA), uric acid (UA), and DA could be separated in the presence of three chemicals. Thereafter, the LOD of the sensor was calculated through DPV analysis, with an LOD of DA of 0.1 nM. Moreover, the rGO-based composite for DA detection was utilised in the cultivation of neuroblastoma cells. In this study, it was demonstrated that the composite was biocompatible, and can identify the EC signals of DA released from living cells through DPV measurement.
Carbon nanotube (CNT)-based dopamine (DA) sensors
According to previous studies, 3D-carbon-based materials have outstanding electrical properties that function advantageously in electrical sensing [72]. For instance, multi-wall carbon nanotubes (MWCNTs), a 3D-carbon-based material with a cylindrical structure, have more than two times higher electrical conductivity than graphene [73]. Through the application of CNTs, Eom et al. reported a novel CNT-based sensing platform for the sensitive detection of DA (Fig. 3a) [54]. To fabricate the sensing platform, the CNTs were modified with overoxidised polypyrrole (OPPy), where its amine group increases the sensitivity and electroactivity of the sensing platform. According to the EC results, the platform exhibits excellent selectivity in the presence of interfering molecules (i.e., ascorbic acid and glucose), and the LOD of the platform was 136 pM (Fig. 3b, c). Additionally, the platform was utilised for the cultivation of dopaminergic neurons. During cultivation, it was confirmed that the platform is biocompatible and a specific amperometric current of DA released from the cells is detectable through EC measurement.
In another study, MWCNTs and AgAu nanoparticle-based electrodes as DA sensors for monitoring DA release have been reported [55]. Interestingly, the MWCNTs used in the sensor were modified with AgAu nanoparticles. The AgAu nanoparticles, as a noble metal, increased the electron transfer rate and enhanced the electrochemical performance of the bare MWCNTs. In addition, the electrochemical sensor was quantified using CV analysis, and its LOD and linear range were 0.23 nM and 3 nM–2.3 mM, respectively. Above all, it was demonstrated that extracellular DA could be monitored in PC12 cells cultured on the sensor. In 2020, a novel MWCNT and graphene quantum dot-based electrochemical sensor to detect extracellular DA was reported [56]. Since the graphene quantum dots used in the modification of MWCNTs increased the surface area of the sensor, the electrical properties of the MWCNTs were improved. Based on the CV results, the LOD of the sensor was calculated as 0.87 nM with a wide linear range of 0.005 to 100 µM. Moreover, the sensor showed good cell biocompatibility through the cell viability test. Above all, the sensor could detect a specific EC DA peak, and the sensor could discriminate the level of DA release induction by KCl.
Taken together, it can be concluded that carbon materials have various advantages in sensing techniques. To be specific, it has been proven that carbon materials possess biocompatible properties and excellent EC features. Furthermore, carbon-materials are readily manufactured using various nanostructures. For these reasons, carbon-materials are outstanding candidates for the detection of DA.
Gold nanostructure-modified electrochemical dopamine sensors
Gold nanoparticles are an advantageous material for biosensing, owing to several properties such as excellent conductivity, signal amplification, and good biocompatibility [74]. In particular, gold nanoparticles have been utilised in various types of biosensors, including localised surface plasmon resonance sensors, optical sensors, and EC sensors [32, 75, 76]. In particular, various EC sensors for the detection of DA released from cells have recently been developed, as gold can enhance cell adhesion and growth and has outstanding electrical properties [77]. In terms of structures, there are three strategies to utilise gold to detect DA released from cells.
Electrochemical dopamine sensors modified with various bare gold structures
In 2019, a new 3D-nanoelectrode modified with gold to detect DA from living cells was devised [58]. In particular, nanoelectrodes have a specific nanocone-shaped 3D structure. To fabricate the gold nanoelectrode, glass capillaries were prepared using laser pulling, and then gold was coated onto the surface of the treated glass electrode. CV confirmed that the gold nanoelectrode showed higher EC peaks toward DA than the bare nanoelectrode. Moreover, the current responses of DA exocytosis from single cells were observed when the gold nanoelectrode was close to the cell.
In 2018, Kim et al. also showed 3D-structural gold nanopillar pattern arrays to detect DA [78]. In this study, laser interference lithography was used to fabricate highly homogeneous polymer nanohole patterns. Subsequently, the EC deposition technique was used to deposit gold into the nanohole patterns, and then, the polymer nanohole patterns were sensitively removed to achieve 3D-structural gold nanopillar pattern arrays. Through CV and amperometry, the platform showed excellent sensing capabilities, with a detection limit of 5.83 µM, and DA was detectable in the presence of interferants. Thus, the gold 3D structure of this study was sensitive to DA because of its high surface area and conductivity. In summary, gold-based 3D structures and highly sensitive DA sensors with high surface areas can be fabricated using electrodeposition techniques. This approach has been utilised to fabricate 3D-structural electrochemical sensors for detection of DA released from living cells in many studies. Recently, a study by Senel et al. used a gold-deposited EC platform on which countless micro-sized pyramid models were fabricated to increase the surface area to be arrayed to effectively detect DA secreted from neuroblastoma cells by EC methods [59]. The sensing electrode was fabricated by incubation, salinisation, and EC deposition, as illustrated in Fig. 4a. The sensing electrode has excellent sensitivity against DA (Fig. 4b). The neuroblastoma cells cultured on the fabricated platform showed excellent potential for the electrocatalytic activity to release DA. The LOD was 0.5 ± 0.01 nM, and the wide linear range was from 10 nM to 500 µM (Fig. 4c). These parameters offer a significant advantage in that real-time analyses directly from living cells can be achieved by measuring the released DA by culturing the neuroblastoma cell on the fabricated platform. Thus, this system also offers a significant advantage owing to its high selectivity and large surface area achieved through surface modification and can recognise the DA secreted from living cells.
Taken together, gold is an outstanding material for the fabrication of DA sensors owing to its high biocompatibility and conductivity, which contribute to the improvement of sensitivity of DA detection from living cells.
Hybrid gold nanoparticle-based electrochemical dopamine sensors
The first strategy is to modify existing electrodes with gold nanoparticles (GNPs), which have been used as an important material for the fabrication of DA sensors owing to several properties, such as good chemical stability and electrical conductivity. Furthermore, GNPs have rich surface chemistry, which can be easily conjugated with other chemicals [79]. Several studies have demonstrated the attractiveness of GNPs as DA sensors. In 2019, a novel gold electrode was modified with GNPs, Nafion™ (NF), and β-cyclodextrin (CD) for EC detection of DA [80]. Specifically, the GNPs were electrodeposited onto the surface of the carbon fibre electrode. According to the cyclic voltammograms, the EC peaks of DA were higher at the GNP-modified gold electrode than at the bare gold electrode, indicating that GNPs functioned as an EC signal enhancer. In addition, the highest EC peaks of DA were obtained at the GNP/NF/CD-modified gold electrode, which showed the synergistic effect of the combination of GNPs and the chemicals on DA sensing. Moreover, DA diluted in urine was detectable using the GNP/NF/CD-modified gold electrode for DPV measurement, which demonstrated that the GNP-based DA sensor can detect DA sensitively in the presence of many interfering molecules. In summary, GNPs are useful materials for the EC detection of DA. In 2018, a carbon fibre electrode, an existing electrode modified with GNPs for detecting DA released from living cells, was reported (Fig. 5a) [57]. To fabricate the GNP-modified carbon fibre electrodes, the GNPs were electrodeposited onto the surface of the carbon fibre electrodes (Fig. 5b). The modified electrodes showed higher EC DA peaks than that of the bare carbon fibre electrode, which indicates that GNPs increased the electron transfer during DA detection and eventually improved the sensitivity of the DA sensor. Furthermore, the levels of DA released from pheochromocytoma cells (PC12) were monitored successfully using the modified electrode through amperometry (Fig. 5c).
Metal oxide-based dopamine (DA) sensors
With the rise of the industrial revolution, researchers have begun to study how to store large amounts of energy [81] efficiently. Batteries made from metal oxide are widely used in mobile phones, laptop computers, among other applications. In particular, metal oxides in nanomaterials are easy to manufacture on a large scale and have highly specific capacities due to abundant redox reactions containing various ions [81]. In this section, we highlighted recent studies using metal oxide nanoparticles and nanostructures as the core material to enhance the EC signals of DA, as well as to improve the biocompatibility of the electrodes.
Morphologically transformed metal oxide-based dopamine (DA) sensors
Specifically, morphologically transformed iron oxide nanoparticles remain an issue for detecting nanomolar units of dopamine under physiological conditions but possess excellent electrolytic properties; hence, they are applied to these iron oxides with electrodes. In 2017, Yang. W. et al. synthesised iron oxides and platinum to create a FePt-Fe3O4 dumbbell-like nanostructure and measured dopamine [60]. The electrode was fabricated by repeated heating and cooling, and the disposal-precipitation processes were repeated. The LOD of the generated platform for DA was 1 nM, and the linear range was 0.1 µM to 90 µM. In addition, selectivity was demonstrated via EC analysis of DA in the presence of 1.0 µM uric acid, 1.0 µM ascorbic acid, and 0.1 µM dopamine. After growing PC12 cells on the platform and stimulating the cells with KCl solution to induce DA release, the electrode recognised released DA, an indication of biocompatibility.
In addition to iron oxide, researchers have investigated other oxides such as nickel oxide, which has excellent electrochemical performance at a low cost to the electrodes. It is environmentally friendly and is well known for its unique and attractive properties as it is used as an excellent electrode material for pseudocapacitors [82]. In 2018, Emran et al. developed a metal oxide nanomaterial platform based on lacy flower-like NiO. The sensor was fabricated by ultrasonication and deposition on an ITO electrode [83]. The detection limit was 85 nM, and the linear range was 0.5 µM to 5 µM. After the PC12 cells were cultured, the supernatant was isolated and measured while simultaneously measuring the isolated supernatant. As the supernatant of PC12 cells was added, the amperometry current was observed to change. This finding indicated that the fabricated sensor has biocompatibility and allows real-time measurement with amperometry because of its ability to measure secreted DA.
Mesoporous metal oxide-based dopamine (DA) sensors
Some metal oxides with generalised structures have the disadvantage of low conductivity [84]. For this reason, research has begun to actively detect analytes by modifying the state of the metal oxide. Typically, attempts are made to transform a metal oxide structure into a mesoporous form because this structure has excellent topological stability, controllability, and veracity. In 2019, an EC platform with a mesoporous design was reported [61]. The scaffolds were fabricated by impregnation, calcination, or etching, and finally, a mesoporous structure was achieved (Fig. 6). Of these electrodes, the best detection limit was 0.8 nM, and the linear range was between 2 and 600 nM. In the presence of ascorbic acid and uric acid, the sensor showed excellent selectivity regardless of the DA detection ability. The DA released from cultured PC12 cells was measured in real-time using amperometry, and changes in current were observed (Fig. 6d). By monitoring the DA released from living cells in real-time using amperometry, these electrodes were shown to be excellent sensors with high biocompatibility, selectivity, and sensitivity.
MFe2O4-based dopamine (DA) sensors
Recently, research has been conducted to create a series of MFe2O4 (M = Ni2+, Mn2+, Fe2+) structures by attaching different metal structures to iron oxide [85]. In particular, ZnFe2O4 has received attention as an excellent material for electrode applications because it is non-toxic, odourless, biocompatible, acid- and alkali-insoluble, and is stable at high temperatures [86]. In 2020, Huang et al. developed a platform based on ZnFe2O4 with mesopores [62]. The platform was fabricated by the hitting method, and showed excellent performance, with a detection limit of 0.4 nM and a linear range of 2–600 nM. The presence of interfering substances such as a tenfold concentration of NaCl, uric acid, and ascorbic acid does not interfere with DA detection, indicating that the fabricated EC platform has excellent selectivity for dopamine. Owing to such excellence in the sensor performance, dopamine exocytosis from PC12 cells was also successfully measured by amperometry in real-time under K+ stimulation condition. To increase the EC detection efficiency, it is critical to add or synthesise a material for application to the electrode.
As a short summary of EC biosensors, the capacity for detecting an analyte varies depending on the electrode structural design or fabrication methods. By applying these features, research is continually progressing, and the sensitivity and selectivity toward the analyte can be achieved with maximum efficiency. Therefore, it is expected that an electrode with excellent detection capacity will be developed soon.