Evaluation of In Vitro Serotonin-Induced Electrochemical Fouling Performance of Boron Doped Diamond Microelectrode Using Fast-Scan Cyclic Voltammetry.

Fast-scan cyclic voltammetry (FSCV) is an electrochemical sensing technique that can be used for neurochemical sensing with high spatiotemporal resolution. Carbon fiber microelectrodes (CFMEs) are traditionally used as FSCV sensors. However, CFMEs are prone to electrochemical fouling caused by oxidative byproducts of repeated serotonin (5-HT) exposure, which makes them less suitable as chronic 5-HT sensors. Our team is developing a boron-doped diamond microelectrode (BDDME) that has previously been shown to be relatively resistant to fouling caused by protein adsorption (biofouling). We sought to determine if this BDDME exhibits resistance to electrochemical fouling, which we explored on electrodes fabricated with either femtosecond laser cutting or physical cleaving. We recorded the oxidation current response after 25 repeated injections of 5-HT in a flow-injection cell and compared the current drop from the first with the last injection. The 5-HT responses were compared with dopamine (DA), a neurochemical that is known to produce minimal fouling oxidative byproducts and has a stable repeated response. Physical cleaving of the BDDME yielded a reduction in fouling due to 5-HT compared with the CFME and the femtosecond laser cut BDDME. However, the femtosecond laser cut BDDME exhibited a large increase in sensitivity over the cleaved BDDME. An extended stability analysis was conducted for all device types following 5-HT fouling tests. This analysis demonstrated an improvement in the long-term stability of boron-doped diamond over CFMEs, as well as a diminishing sensitivity of the laser-cut BDDME over time. This work reports the electrochemical fouling performance of the BDDME when it is repeatedly exposed to DA or 5-HT, which informs the development of a chronic, diamond-based electrochemical sensor for long-term neurotransmitter measurements in vivo.

In Vitro Biofouling Performance of Boron-Doped Diamond Microelectrodes for Serotonin Detection Using Fast-Scan Cyclic Voltammetry.

Neurotransmitter release is important to study in order to better understand neurological diseases and treatment approaches. Serotonin is a neurotransmitter known to play key roles in the etiology of neuropsychiatric disorders. Fast-scan cyclic voltammetry (FSCV) has enabled the detection of neurochemicals, including serotonin, on a sub-second timescale via the well-established carbon fiber microelectrode (CFME). However, poor chronic stability and biofouling, i.e., the adsorption of interferent proteins to the electrode surface upon implantation, pose challenges in the natural physiological environment. We have recently developed a uniquely designed, freestanding, all-diamond boron-doped diamond microelectrode (BDDME) for electrochemical measurements. Key potential advantages of the device include customizable electrode site layouts, a wider working potential window, improved stability, and resistance to biofouling. Here, we present a first report on the electrochemical behavior of the BDDME in comparison with CFME by investigating in vitro serotonin (5-HT) responses with varying FSCV waveform parameters and biofouling conditions. While the CFME delivered lower limits of detection, we also found that BDDMEs showed more sustained 5-HT responses to increasing or changing FSCV waveform-switching potential and frequency, as well as to higher analyte concentrations. Biofouling-induced current reductions were significantly less pronounced at the BDDME when using a "Jackson" waveform compared to CFMEs. These findings are important steps towards the development and optimization of the BDDME as a chronically implanted biosensor for in vivo neurotransmitter detection.

Boron doped diamond thin films for the electrochemical detection of SARS-CoV-2 S1 protein.

Amidst a global pandemic, a precise and widely accessible rapid detection method is needed for accurate diagnosis and contact tracing. The lack of this technology was exposed through the outbreak of SARS-CoV-2 beginning in 2019. This study sets the foundation for the development of a boron doped diamond (BDD)-based impedimetric sensor. While specifically developed for use in the detection of SARS-CoV-2, this technology uses principles that could be adapted to detect other viruses in the future. Boron doped polycrystalline diamond electrodes were functionalized with a biotin-streptavidin linker complex and biotinylated anti-SARS-CoV-2 S1 antibodies. Electrodes were then incubated with the S1 subunit of the SARS-CoV-2 spike surface protein, and an electrical response was recorded using the changes to the electrode's charge transfer resistance (Rct), measured through electrochemical impedance spectroscopy (EIS). Detectable changes in the Rct were observed after 5-min incubation periods with S1 subunit concentrations as low as 1 fg/mL. Incubation with Influenza-B Hemagglutinin protein resulted in minimal change to the Rct, indicating specificity of the BDD electrode for the S1 subunit of SARS-CoV-2. Detection of the S1 subunit in a complex (cell culture) medium was also demonstrated by modifying the EIS protocol to minimize the effects of sample matrix binding. BDD films of varying surface morphologies were investigated, and material characterization was used to give insight into the microstructure-performance relationship of the BDD sensing surface.

Next-Generation Diamond Electrodes for Neurochemical Sensing: Challenges and Opportunities.

Carbon-based electrodes combined with fast-scan cyclic voltammetry (FSCV) enable neurochemical sensing with high spatiotemporal resolution and sensitivity. While their attractive electrochemical and conductive properties have established a long history of use in the detection of neurotransmitters both in vitro and in vivo, carbon fiber microelectrodes (CFMEs) also have limitations in their fabrication, flexibility, and chronic stability. Diamond is a form of carbon with a more rigid bonding structure (sp3-hybridized) which can become conductive when boron-doped. Boron-doped diamond (BDD) is characterized by an extremely wide potential window, low background current, and good biocompatibility. Additionally, methods for processing and patterning diamond allow for high-throughput batch fabrication and customization of electrode arrays with unique architectures. While tradeoffs in sensitivity can undermine the advantages of BDD as a neurochemical sensor, there are numerous untapped opportunities to further improve performance, including anodic pretreatment, or optimization of the FSCV waveform, instrumentation, sp2/sp3 character, doping, surface characteristics, and signal processing. Here, we review the state-of-the-art in diamond electrodes for neurochemical sensing and discuss potential opportunities for future advancements of the technology. We highlight our team's progress with the development of an all-diamond fiber ultramicroelectrode as a novel approach to advance the performance and applications of diamond-based neurochemical sensors.

Flexible, diamond-based microelectrodes fabricated using the diamond growth side for neural sensing.

Diamond possesses many favorable properties for biochemical sensors, including biocompatibility, chemical inertness, resistance to biofouling, an extremely wide potential window, and low double-layer capacitance. The hardness of diamond, however, has hindered its applications in neural implants due to the mechanical property mismatch between diamond and soft nervous tissues. Here, we present a flexible, diamond-based microelectrode probe consisting of multichannel boron-doped polycrystalline diamond (BDD) microelectrodes on a soft Parylene C substrate. We developed and optimized a wafer-scale fabrication approach that allows the use of the growth side of the BDD thin film as the sensing surface. Compared to the nucleation surface, the BDD growth side exhibited a rougher morphology, a higher sp 3 content, a wider water potential window, and a lower background current. The dopamine (DA) sensing capability of the BDD growth surface electrodes was validated in a 1.0 mM DA solution, which shows better sensitivity and stability than the BDD nucleation surface electrodes. The results of these comparative studies suggest that using the BDD growth surface for making implantable microelectrodes has significant advantages in terms of the sensitivity, selectivity, and stability of a neural implant. Furthermore, we validated the functionality of the BDD growth side electrodes for neural recordings both in vitro and in vivo. The biocompatibility of the microcrystalline diamond film was also assessed in vitro using rat cortical neuron cultures.

Isatin Detection Using a Boron-Doped Diamond 3-in-1 Sensing Platform.

Boron-doped diamond (BDD) is a promising electrochemical tool that exhibits excellent chemical sensitivity and stability. These intrinsic advantages coupled with the material's vast microfabrication flexibility make BDD an attractive sensing device. In this study, two different 3-in-1 BDD electrode sensors were fabricated, characterized, and investigated for their capability to detect isatin, an anxiogenic indole that possesses anticonvulsant activity. Each device was comprised of a working, reference, and auxiliary electrode, all made of BDD. Two different working electrode geometries were studied, a 2 mm diameter macroelectrode (MAC) and a microelectrode array (MEA). The BDD quasi-reference electrode was studied by measuring its potential against a traditional Ag/AgCl reference electrode. While the potential shifted as a function of solution pH, a miniscule potential drift was observed when holding the solution pH constant. Specifically, the BDD quasi-reference electrode had a potential of -0.2 V (vs Ag/AgCl) in a pH 7 solution, and this remained stable for a 30-h time period. For the detection of isatin, solutions were analyzed using both sensors in pH 7.4 phosphate buffered saline (PBS). Using the MEA sensor, the limit of detection (LOD, (3σ)/m) for isatin was found to be 0.04 µM; an increase to 0.22 µM was observed with the MAC sensor. These results were compared to those obtained from UV-vis spectrophotometry, where a 0.57 µM LOD was observed. The feasibility for use in a complex sample matrix was also examined by completing measurements in urine simulant. The results presented herein indicate that both 3-in-1 BDD sensors are applicable at low limits of detection with potential application as an electrochemical detector for chromatographic methods.

Large-scale, all polycrystalline diamond structures transferred onto flexible Parylene-C films for neurotransmitter sensing.

Boron-doped diamond (BDD) has superior electrochemical properties for bioelectronic systems. However, due to its high synthesis temperature, traditional microfabrication methods have limits to integrating BDD with emerging classes of flexible, polymer-based bioelectronic systems. This paper introduces a novel fabrication solution to this challenge, which features (i) a wafer-scale substrate transfer process with all diamond structures transferred onto a flexible Parylene-C substrate and (ii) Parylene anchors introduced to strengthen the bonding between BDD and Parylene substrates, as demonstrated by a peeling test. The electrochemical properties of the transferred BDD-polymer electrodes are evaluated using (i) an outer sphere redox couple Ru(NH3)62+/3+ to study the electron transfer process and (ii) quantitative and qualitative studies of the neurotransmitter redox couple dopamine/dopamine-o-quinone. A linear response of the BDD sensor to dopamine concentrations of 0.5 µM to 100 µM is observed (R2 = 0.999) with a sensitivity of 0.21 µA cm-2 µM-1. These examples of fabricated diamond-polymer devices suggest a broad application in advanced bioelectronics and optoelectronics.