Translating Extracellular Electron Transfer Activities with Organic Electrochemical Transistors

Summary

Here, we present a protocol for using organic electrochemical transistors (OECTs) to translate extracellular electron transfer (EET) activity in Shewanella oneidensis into electrical signals. The hybrid OECT system provides enhanced robustness, sensitivity, and potential for rapid, high-throughput testing, making it an effective tool for EET measurements.

Abstract

Extracellular electron transfer (EET) is a process through which certain microorganisms can transfer electrons across their cell membranes to external electron acceptors, linking cellular metabolism to their environment. While Geobacter and Shewanella have been the primary models for EET research, emerging studies reveal that EET-active species are also associated with fermentation and the human gut microbiome. Leveraging the ability of EET to bridge biological and electronic systems, we present a protocol for using organic electrochemical transistors (OECTs) to translate microbial EET activity into easily detectable electrical signals. This system enables the use of cellular responses to external stimuli for biosensing and biocomputing applications. Specifically, we demonstrated the de-doping of the p-type poly(3,4-ethylenedioxythiophene): poly(styrenesulfonate) (PEDOT: PSS) channel in the OECT is driven by cellular EET from Shewanella oneidensis. By transcriptionally controlling EET flux by genetic circuits, we establish the biosensing capability of this hybrid OECT system to detect chemical stimuli, such as inducer molecules. Furthermore, we introduce plasmid-based Boolean logic gates within the cells, allowing them to process environmental signals and drive current changes in the OECTs, further demonstrating the biocomputing potential of these devices. This method provides a novel interface between biological systems and electronics, enabling future high-throughput screening, biosensing, and biocomputing applications.

Introduction

Devices that can transduce and amplify biological and chemical activities into electrical signals are crucial across various fields, such as sensing^1,2, neuromorphic computing^3,4, and wearable electronics⁵. Among these, organic electrochemical transistors (OECTs) have emerged as exceptional interfaces between biological systems and electronic readouts due to their compatibility with aqueous environments and low operating voltages^6,7. OECTs differ from conventional electr....

Protocol

NOTE: All chemicals were used as received without further purification. If not specified, analytical-grade chemicals were used.

1. OECT device fabrication

NOTE: The OECTs are fabricated on quartz microscope slides using standard microfabrication techniques adapted from prior work²⁴. As shown in Figure 2A, eight OECT devices are arranged on a single standard slide. Pre-cut polydimethylsiloxane (PDMS) sheets are placed on the slide to form OECT chambers and access ports. The p-type conducting polymer poly(3,4-ethylenedioxythiophene): poly(styrenes....

Results

Fitted rate constant k
The fitted rate constant k of the OECT channel current I_DS serves as a reliable metric for assessing the EET activity of the sample. While the constant gate bias voltages would affect the rate constants, we choose 0.2 V to ensure positive bias to encourage bacterial electron transfer while avoiding fast de-doping at higher gate voltages and providing and minimizing electrochemical stress on the bacteria cells²³

Discussion

Electrochemical Cell (EC) Comparison
One major advantage of fitting the rate constant from the OECT channel current is that it minimizes device variation by focusing on the underlying dynamics of I_DS changes rather than the raw output. Combined with the inherent signal amplification capability of OECTs, this approach enhances the robustness of hybrid OECT systems compared to traditional electrochemical cells (EC). For example, at the end of each 24-hour OECT experiment, transfer curves a.......

Materials

Name	Company	Catalog Number	Comments
3-oxohexanoyl-homoserine lactone	Sigma-Aldrich
anhydrotetracycline hydrochloride	VWR
casamino acids	VWR
Equipment
Ethylene glycol	Sigma-Aldrich		anhydrous 99.8%,
HEPES buffer solution	VWR		1 M in water, pH = 7.3
isopropyl ß-D-1-thiogalactopyranoside	Teknova
kanamycin sulfate	Growcells
Magnesium(II) sulfate heptahydrate	VWR
PEDOT:PSS aqueous suspension	Heraeus Epurio LLC	Clevios PH1000
Potassium phosphate dibasic	Sigma-Aldrich
Potassium phosphate monobasic	Sigma-Aldrich
Potentiostat	PalmSens BV	MultiPalmSens4
Quartz microscopic slides	AdValue	FQ-S-003
Quartz microscopic slides
Sodium chloride	VWR
Sodium DL-lactate	TCI		60% in water
Sodium fumarate	VWR		98%
Sulfuric acid	Sigma-Aldrich		95.0%-98.0%
Two-part silicone elastomer	Electron Microscopy Sciences	Sylgard184
Wolfe's Trace Mineral Mix	ATCC