Electrochemical Sensing for Neuropeptide-Y Measurements

Abstract

Neuropeptide Y (NPY), the most abundant neuropeptide in the human body, plays a pivotal role in regulating stress, appetite, and neurological processes, making its precise quantification essential for understanding its contributions to disorders like obesity, post-traumatic stress disorder (PTSD), and anxiety. This thesis presents the development and characterization of electrochemical biosensors for the selective and sensitive detection of NPY, using aptamer-modified and molecularly imprinted polymer (MIP)-based microelectrodes with electrochemical impedance spectroscopy (EIS). The research addresses key challenges in neurochemical sensing, including biofouling, selectivity in complex biological matrices, and the need for high spatial and temporal resolution. Aptamer-modified microelectrodes, fabricated on carbon fiber and platinum substrates, were optimized to mitigate biofouling and enhance NPY detection. Comparative studies revealed that platinum microelectrodes, when functionalized with aptamers, exhibit fouling characteristics comparable to carbon fiber, broadening material options for biosensor design. Using EIS, we measured changes in the imaginary impedance component (-ω*Zimag), which correlates with surface capacitance and NPY adsorption, achieving a linear detection range of 10 ng/mL to 1,000 ng/mL in artificial cerebrospinal fluid without redox probes. Label-free detection on planar aptamer-modified electrodes further improved sensitivity, detecting NPY at concentrations as low as 20 pg/mL, with a linear range from 20 pg/mL to 1,000 pg/mL. The adsorption process followed the Langmuir isotherm model, contrasting with the Freundlich model observed for microelectrodes, providing a robust framework for quantification. Selectivity was confirmed against interfering molecules, including dopamine, serotonin, and structurally similar peptides, even at 20-fold higher concentrations, underscoring the specificity of aptamer-based recognition. In parallel, molecularly imprinted polypyrrole (MIPpy) was electrodeposited on carbon fiber microelectrodes to create NPY-specific binding sites. CV and EIS measurements revealed linear correlations between oxidation current, capacitance, and NPY concentrations (50 ng/mL to 1,000 ng/mL), demonstrating the potential of MIPpy-based sensors for high-resolution brain measurements. Incorporating methylene blue as a redox probe enhanced detection sensitivity to 5 pg/mL, with optimal performance at low frequencies and potential aligned with the probe’s redox characteristics. These biosensors offer exceptional temporal and spatial resolution, making them promising in vivo NPY monitoring tools. Challenges such as non-specific protein adsorption in biological fluids were identified, suggesting that advanced surface passivation strategies were needed. This work establishes a foundation for real-time neurochemical sensing, with applications in studying NPY dynamics and developing diagnostic tools for neurological disorders. Future research will focus on optimizing sensor designs for in vivo use and extending the approach to other neuropeptides, advancing the field of electrochemical biosensing.