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Table · dataset · 2026

Diffusion-controlled Third-generation Electrochemical NADH Biosensor with the Diaphorase Subunit of Formate Dehydrogenase 1 (Supporting Information)

Listed in ZivaHub and Deakin Research Online and DMU Figshare and HKU DataHub and Swinburne Figshare and DaYta Ya Rona and SUNScholarData and figshare and Loughborough Research Repository and GRANTS Data and UP Research Data Repository — shown once because both records carry DOI 10.50892/data.electrochemistry.33719344.v1

<p dir="ltr">Nicotinamide adenine dinucleotide (NAD) serves as a crucial biomarker for cellular metabolism and various diseases.

Description

In this study, we present a third-generation (3G) electrochemical biosensor for NADH that operates under diffusion control and is founded on direct electron transfer (DET)-type bioelectrocatalysis. The biosensor was developed utilizing the diaphorase subunit of formate dehydrogenase 1 expressed in <i>Escherichia coli</i> (EcFoDH1B) as the biocatalyst.

To achieve diffusion-controlled sensing, we engineered the enzyme-electrode interface through thiol modification of microporous gold electrodes to enhance the DET between EcFoDH1B and the electrode. The resultant biosensor, utilizing a 100 µm-diameter microelectrode, displayed a linear response to NADH concentrations up to 750 µM, with the calibration curve slope closely matching the theoretical value predicted from hemispherical diffusion.

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The biosensor exhibited high linearity (the coefficients of determination (<i>R</i><sup>2</sup> values) > 0.999), sensitivity (120 ± 10 µA mM<sup>−1</sup> cm<sup>−2</sup>), selectivity against NADPH at 100 µM, and rapid response, achieving a steady-state current within 50 s. These findings showcase the first diffusion-controlled 3G NADH biosensor and offer a design approach for highly reproducible and potentially calibration-free electrochemical biosensing based on DET-type bioelectrocatalysis.</p>

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Biochemistry and cell biology 71%
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