Development of an In Vivo, Real-Time and Continuous Insulin Sensor Utilizing an Extended Gate Field Effect Transistor-Based Transducer
Authors: Khanwalker Mukund , Hatada Mika , Wilson Ellie , Probst David , Twiddy Jack , Daniele Michael , Asano Ryutaro , Ikebukuro Kazunori , Sode Koji 
Published: December 2024

Introduction

In this paper, we aim to utilize an extended gate field-effect transistor (EGFET)-based transducer to measure insulin. The design of EGFETs is characterized by a unique structural arrangement, where a physical barrier exists between the semiconductor component and the analyte or physical medium. Due to this structure, EGFET possesses the advantage of protecting the electrode from intricacies within the measurement solution or medium. The need for a redox probe is also effectively eliminated by incorporating EGFETs, marking a significant breakthrough in electrochemical measurement. The lack of a redox probe allows EGFET-based sensors to conduct measurements with minimal invasiveness and enhanced biocompatibility in accordance with an in vivo monitoring system. As the target analyte is not required to be electrochemically active, EGFET is highly customizable for a wide range of targets, including neurotransmitters, biomarkers, and metabolites. Therefore, we constructed immunosensors immobilized with anti-insulin single-chain antibody (scFv) and measured insulin levels, with the objective of creating an in vivo sensor designed to improve glycemic management. We aimed to implement the inherent semiconductor properties of EGFETs by performing real-time monitoring of changes in electrical properties to quantify various insulin concentrations.

Methods

Development of an in vivo sensor requires the use of a platform that can be implemented within an in vivo environment. For this purpose, we utilized a gold microwire electrode immobilized with anti-insulin scFv. The gold microwire electrodes were prepared by electrochemically cleaning and further roughening them to increase the active surface area for the purpose of improving immobilization of the antibody. To achieve site-specific immobilization of anti-insulin scFv, we employed a nickel-chelated nitrilotriacetate self-assembled monolayer (Ni-NTA SAM). As the scFv harbors a histidine tag and Ni-NTA has an affinity for histidine residues, we were able to achieve oriented immobilization on the electrode surface. Within this study, we compared the effects of covalent immobilization, which leads to random orientation with the antibody, and non-covalent oriented immobilization by comparing the signal obtained from EGFET measurement. EGFET measurements were conducted using a transistor consisting of source, drain, and gate terminals. In this configuration, a constant potential was maintained between the source and drain terminals, while a sweeping potential was applied between an Ag/AgCl reference electrode and the source terminal. Measurement of insulin was performed within a buffer and artificial interstitial fluid (aISF) to model an in vivo environment.

Results and Discussion

The interaction between insulin and the immobilized anti-insulin scFv on the electrode surface induces a shift in surface potential, driven by the redistribution of charges across the extended gate. The change in surface potential leads to a subsequent modification of the gate potential, known as Vg. The gate potential, when less than the threshold voltage (Vth), operates in the cutoff region where little to no current is present. However, when Vg is greater than Vth, the system operates in the ohmic region where current can flow. The following relationship allowed us to monitor the drain-source current or Ids within the ohmic region to evaluate insulin concentrations. The signal change due to insulin binding to anti-insulin scFv was observed at a reference voltage of 0.6 volts.

Conclusion

We measured insulin concentrations using an electrode modified with anti-insulin scFv and demonstrated that EGFET can be used as a working principle for an in vivo monitoring system. Additionally, the measurement principle does not require a redox probe within the measurement solution. Future work will include implanting the anti-insulin scFv immobilized gold microwire electrode within a rat model and measuring insulin.