High resolution voltammetric and field-effect transistor readout of carbon fiber microelectrode biosensors

Rapid and sensitive pH measurements with increased spatiotemporal resolution are imperative to probe neurochemical signals and illuminate brain function. We interfaced carbon fiber microelectrode (CFME) sensors with both fast scan cyclic voltammetry (FSCV) and field-effect transistor (FET) transducers for dynamic pH measurements. The electrochemical oxidation and reduction of functional groups on the surface of CFMEs affect their response over a physiologically relevant pH range. When measured with FET transducers, the sensitivity of the measurements over the measured pH range was found to be (101 ± 18) mV, which exceeded the Nernstian value of 59 mV by approximately 70%. Finally, we validated the functionality of CFMEs as pH sensors with FSCV ex vivo in rat brain coronal slices with exogenously applied solutions of varying pH values indicating that potential in vivo study is feasible.

KCl solution, protruding CFME tips were epoxied and then rinsed in acetone to wash away any excess residual epoxy. The electrodes were cured in the oven for 4 h at 125 °C.

Scanning Electron Microscope (SEM). The thickness and morphologies of CFMEs were
obtained with a JEOL JSM-IT100 (JEOL, Tokyo, Japan) at a 10 kV accelerating voltage.
Field Effect Transistor (FET). We fabricated a commercially sourced single-gate FET combined with signal processing to significantly improve the overall performance of the complete sensor system. Here, lock-in amplifier (LIA) was widely used to improve the overall signal-to-noise ratio (SNR) by allowing the recovery of weak signals at a specific reference frequency and phase. The pH resolution of FET can be further increased by integrating LIA to recover weak signals. For the measurement, CFMEs applied to the top gate modulate the current in the semiconducting channel of FET. An Ag/AgCl reference electrode was connected to the output of PID (proportionalintegral-derivative) controller, and it was used to adjust the gate potential and maintain a constant channel current. PID controllers are widely used to control process variables for increased stability and accuracy by continually adjusting a process parameter in response to deviations from a predetermined set point. The PID is commonly used in controlled temperature sensors, gas detectors, photosensors, and hydrogen sensors. Moreover, it is also applied to improve the reproducibility and accuracy of atomic force microscopy (AFM) by controlling gain parameters.

Fast-Scan Cyclic
where QH2s and Qs indicate the reduced and oxidized form of a surface hydroquinone-like moiety.
QH-peak shows the transition for the hydroquinone to quinone on the forward scans while Q-peak indicates the transition for the quinone to hydroquinone on the backward scans. 3

Tissue Extraction. Tissue samples were collected from female Sprague Dawley rats in accordance
with IACUC and animal facility protocols at American University (Protocol #20-09). Rats were housed in 12 hour light and dark cycles and provided food and water ad libidum. Firstly, a rat was removed from the cage and placed in a CO2 euthanasia chamber. Reflexes of a rat were then assessed via toe and tail pinch, euthanasia was subsequently confirmed by cervical dislocation.
The head was decapitated using surgical scissors and the skull was exposed by removing surrounding tissue. Large, surgical rongeurs were used to quickly peel away the skull bones to expose and remove the brain. The excised brain was placed into a vial filled with artificial cerebrospinal fluid (aCSF) and stored on ice until use. For slice preparation, multiple coronal cuts were made to target either the caudate putamen or the hippocampus. Coordinates and anatomical landmarks were located according to the Paxinos Rat Brain Atlas. 4 Brain tissue preparation procedures were adapted from Papouin and Haydon. 5 The brain slice was placed into the well of a 24-well plate (Castar, Corning, New York). The slice was then saturated with a cold aCSF buffer, which had been oxygenated by bubbling carbogen gas (95 % O2, 5 % CO2) using an air stone and airline tubing. Subsequently, CFMEs was lowered until it made a contact with the brain tissue and was allowed to equilibrate at least for 15 min with a triangle waveform. aCSF buffer solutions having different pH ranged from 2 to 8 were then exogenously applied by injecting 250 µL of each solution into the slice and adjacent to the CFMEs.
After each injection, the residual aCSF solution in the 24-well plate was discarded and a fresh, oxygenated aCSF was added. Injections were repeated 3 times at each pH with 10 min intervals between them.