2026-10-14T08:55:39Z https://api.figshare.com/v2/oai

oai:figshare.com:article/33719344 2026-10-01T01:17:17Z category_43791 portal_791 item_type_3 month_year_10_2026

Diffusion-controlled Third-generation Electrochemical NADH Biosensor with the Diaphorase Subunit of Formate Dehydrogenase 1 (Supporting Information) Eole FUKAWA (24868586) Taiki ADACHI (24868589) Yuki KITAZUMI (11181993) Osamu SHIRAI (24868592) Keisei SOWA (24868616) Chemistry Direct Electron Transfer-type Bioelectrocatalysis Third-generation Biosensor Electrochemical NADH Sensor Microelectrode <p dir="ltr">Nicotinamide adenine dinucleotide (NAD) serves as a crucial biomarker for cellular metabolism and various diseases. 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. The biosensor exhibited high linearity (the coefficients of determination (<i>R</i>² values) > 0.999), sensitivity (120 ± 10 µA mM⁻¹ cm⁻²), 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> 2026-10-01T00:00:00Z Dataset Dataset 10.50892/data.electrochemistry.33719344.v1 https://figshare.com/articles/dataset/Diffusion-controlled_Third-generation_Electrochemical_NADH_Biosensor_with_the_Diaphorase_Subunit_of_Formate_Dehydrogenase_1_Supporting_Information_/33719344 CC BY 4.0