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

At a Glance

Metadata	Details
Publication Date	2023-05-25
Journal	Biosensors
Authors	Bhavna Gupta, Mason L. Perillo, James R. Siegenthaler, Isabelle E. Christensen, Matthew P. Welch
Institutions	Michigan State University, Fraunhofer USA
Citations	14
Analysis	Full AI Review Included

Executive Summary

Core Value Proposition: Freestanding, all-diamond Boron-Doped Diamond Microelectrodes (BDDMEs) offer significantly improved stability and resistance to biofouling compared to traditional Carbon Fiber Microelectrodes (CFMEs) for chronic in vivo neurotransmitter detection.
Biofouling Resistance: BDDMEs exhibited superior resilience to protein adsorption (BSA soak model). Using the 5-HT-specific Jackson waveform, BDDMEs showed an average current decrease of only 39.01%, significantly less than the 62.5% decrease observed on CFMEs.
Electrochemical Stability: BDDMEs maintained stable anodic peak responses across varying switching potentials (1.0 V to 1.5 V) and demonstrated higher resilience to increasing waveform application frequencies (up to 100 Hz).
Concentration Range: BDDMEs maintained an increasing, non-linear current response up to 100 µM 5-HT, whereas CFMEs saturated and decreased in response above 25 µM, suggesting better resistance to byproduct fouling at high analyte concentrations.
Sensitivity Trade-off: CFMEs delivered superior initial sensitivity and lower Limits of Detection (LODs) (0.049 µM standard waveform) compared to BDDMEs (0.26 µM standard waveform), primarily due to the BDDME’s smaller electroactive area (100-200 µm² vs. 1138-1578 µm² for CFMEs).
Detection Mechanism: The BDDME response suggests a dual modality or diffusion-controlled process, while the CFME response is characteristic of adsorption-controlled kinetics.

Technical Specifications

Parameter	Value	Unit	Context
BDD Growth Method	Microwave CVD (915 MHz)	N/A	Wafer fabrication
BDD Growth Temperature	900	°C	Synthesis condition
BDD Growth Pressure	60	Torr	Synthesis condition
B/C Doping Ratio	37,500	ppm	Boron concentration
BDD Thickness	2-4	µm	Electroactive layer thickness
BDDME Electroactive Area	100 to 200	µm²	Based on 50 µm wide pattern
CFME Diameter	7.4	µm	Unsized AS4 carbon fiber
CFME Exposed Length	100-150	µm	Estimated geometric area: 1138 to 1578 µm²
Standard Waveform Scan Rate	400	V s^-1	Triangular sweep (-0.4 V to 1.3 V)
Jackson Waveform Scan Rate	1000	V s^-1	N-shaped sweep (0.2 V to 1.0 V to -0.1 V)
CFME LOD (Standard WF, Pre-Fouling)	0.049	µM	5-HT detection
BDDME LOD (Standard WF, Pre-Fouling)	0.26	µM	5-HT detection
CFME Current Decrease (Jackson WF)	-62.5	%	Average decrease after BSA biofouling
BDDME Current Decrease (Jackson WF)	-39.01	%	Average decrease after BSA biofouling
BSA Soak Concentration	40	g L^-1	Biofouling model (4% BSA in aCSF)

Key Methodologies

BDDME Wafer Fabrication: BDD films were grown on 4-inch silicon wafers using 915 MHz Microwave CVD at 900 °C and 60 Torr, utilizing 2% CH₄ and a B/C ratio of 37,500 ppm.
Electrode Patterning and Release: Electrodes were patterned via photolithography and RIE etching. They were released from the silicon substrate using an HF:HNO₃:CH₃COOH etchant (5:11:6).
PCD Insulation: The freestanding BDDMEs were fully insulated with Polycrystalline Microcrystalline Diamond (PCD) grown via Hot Filament CVD (HF-CVD) at 35 Torr and 2% CH₄.
CFME Fabrication: CFMEs were hand-fabricated by aspirating 7.4 µm Ø AS4 carbon fibers into glass capillaries, pulled using a micropipette puller, and cut to an exposed length of 100-150 µm.
FSCV Instrumentation: Experiments utilized a two-electrode setup (working electrode vs. quasi Ag/AgCl reference) in a custom flow injection cell (0.75 mL min^-1 flow rate). Data acquisition used an NI-6363 card and HDCV software.
Waveform Testing: Both the standard triangular waveform (-0.4 V to 1.3 V, 400 V s^-1, 10 Hz) and the N-shaped Jackson waveform (0.2 V to 1.0 V to -0.1 V, 1000 V s^-1, 10 Hz) were tested across varying scan rates, holding potentials, switching potentials, and frequencies.
Biofouling Protocol: Electrodes were pre-calibrated, soaked for 12-14 hours in 4% Bovine Serum Albumin (BSA) solution in aCSF (pH 7.4), and then post-calibrated to quantify the reduction in 5-HT oxidative current and sensitivity.

Commercial Applications

The development of microfabricated, freestanding Boron-Doped Diamond Microelectrodes (BDDMEs) is highly relevant to several high-tech sectors:

Chronic Neurochemical Sensing: BDDMEs are optimized for long-term implantation, offering a pathway for chronic monitoring of neurotransmitters (Serotonin, Dopamine) in the brain, essential for research into psychiatric and neurological disorders.
Biocompatible Implants: The all-diamond structure provides superior mechanical and chemical stability, reducing the immune response (gliosis) and biofouling associated with traditional polymer-insulated electrodes.
High-Stability Biosensors: The resistance of BDD to fouling and its wide working potential window make it ideal for biosensors operating in complex, protein-rich biological fluids or harsh chemical environments.
Wafer-Scale Microfabrication: The use of wafer processing allows for batch fabrication of highly consistent, customizable electrode geometries, enabling high-throughput manufacturing and complex array designs for neural interfaces.
Advanced Electrochemical Detection: BDD technology is applicable beyond neuroscience, including high-sensitivity detection of electroactive compounds in pharmaceutical analysis, environmental monitoring, and industrial quality control where electrode stability is paramount.

View Original Abstract

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.

Tech Support

Original Source

DOI: https://doi.org/10.3390/bios13060576

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