MWCNTs-COOH/Nafion modified PCB gold electrode for electrochemical detection of dopamine secreted by SKPs-derived dopaminergic neurons

Low-cost PCB‑gold electrode with superior electron kinetics. • MWCNTs/Nafion network boosts sensitivity and selectivity. • High performance: 1–70 μM linear range and 202 nM LOD. • Strong anti-biofouling and selectivity in complex media. • Direct dopamine detection in 3D dECM neuronal models. Dopamine (DA) is a crucial neurotransmitter whose abnormal fluctuations are linked to neurological disorders such as Parkinson's disease. Detecting DA in complex biological matrices faces challenges from coexisting interferents and electrode biofouling. Here, we developed a carboxylated multi-walled carbon nanotubes (MWCNTs-COOH) and Nafion-modified printed circuit board gold electrode (PCB-AuE) for selective DA detection. The PCB-AuE platform offers superior electron transfer kinetics, ultra-low cost, and design flexibility compared to commercial electrodes. The synergistic MWCNTs-COOH/Nafion modification provides a highly conductive 3D network with abundant active sites while electrostatically repelling anionic interferents (ascorbic acid and uric acid). The sensor achieved a broad linear range (1–70 μM), high sensitivity (1.98 μA/μM), and low detection limit (202 nM) with excellent reproducibility (RSD = 4.8%). Despite an 8.5% signal attenuation in culture media versus PBS, the sensor maintained reliable detection capability. The platform successfully detected endogenous DA secreted by skin-derived precursors (SKPs)-derived dopaminergic neurons cultured in 3D brain decellularized extracellular matrix scaffolds. This disposable, cost-effective sensor provides a practical analytical tool for neurotransmitter monitoring in complex 3D tissue models with significant potential for disease diagnosis and drug screening applications. Schematic representation of the fabrication and application of a nanocomposite-modified PCB gold electrode array for dopamine detection in 3D neuronal models. Sensor Fabrication and Surface Modification. A Printed Circuit Board (PCB) gold electrode array is utilized as the transducer platform. The three-electrode system consists of a central Working Electrode (WE), a surrounding Counter Electrode (CE), and a Pseudo-Reference Electrode (Pseudo-RE). The WE surface is modified via drop-casting with a nanocomposite ink comprising carboxylated multi-walled carbon nanotubes (MWCNTs-COOH) and Nafion. The MWCNTs-COOH provide a high-surface-area conductive network, while Nafion acts as a cation-exchange polymer. Construction of the 3D Dopaminergic Neuronal Model. Skin-derived precursor cells (SKPs) are cultured as spheres and subjected to a 14-day neural induction protocol to generate mature dopaminergic neurons (SKPs-DA), confirmed by tyrosine hydroxylase (TH + ) expression. These neurons are subsequently encapsulated within a decellularized extracellular matrix (dECM) scaffold to create a biomimetic 3D SKPs-DA neuronal tissue. Cell viability within the scaffold is validated via Live/Dead staining (Calcein-AM/Ethidium homodimer-1). Electrochemical Dopamine (DA) Detection and Sensing Mechanism. The modified electrode exhibits high sensitivity (1.98 μA/μM) and a low limit of detection (202 nM) within a linear range of 1–70 μM. The sensing mechanism leverages the synergistic effects of the nanocomposite: (1) The negatively charged sulfonate groups in Nafion electrostatically attract cationic dopamine (DA + ). (2) Anionic species such as ascorbic acid (AA) and uric acid (UA) are repelled by the Nafion layer, enhancing selectivity. (3) Differential Pulse Voltammetry (DPV) shows a distinct oxidation peak at approximately 0.1 V for the SKPs-DA supernatant compared to undifferentiated controls, demonstrating the sensor's efficacy in monitoring neurotransmitter release in complex biological matrices. Download: Download high-res image (125KB) Download: Download full-size image