<p>Electrochemical sensing is a promising approach for the selective and highly sensitive sensing of oxidized and reduced states of Nicotinamide adenine dinucleotide (NAD⁺/NADH). However, the limited selectivity, large overpotential requirements and the electrode fouling concerns associated with the till date reported NADH-specific electrodes continue to impede their potential utility for the design and development of fast, inexpensive and highly reliable point of care devices for electrochemical sensing of NAD⁺ and NADH. Herein we present a simple covalent functionalization approach for the design and development of Cytochrome-c (Cyt-c) functionalized Cu-BDC MOF (Cyt-c/Cu-BDC) as a novel Cu-Fe based bio-mimic for electrochemical sensing of NADH. Our detailed physical, chemical and electrochemical investigations carried out over the so designed Cyt-c/Cu-BDC composite establish it as an electronically conducting, electrochemically stable redox-active electrode material with an exceptional activity towards the selective and ultrasensitive electrochemical sensing of NADH. We demonstrate the practical utility of Cyt-c/Cu-BDC composite for accurate and sensitive electrochemical sensing of NADH in the pico-molar concentration range. The herein demonstrated extremely low LOD (10.4&#xa0;pM), high sensitivity (12.02 ± 0.119 μA nM⁻¹&#xa0;cm⁻²), good anti-interference ability and prolonged stability of the Cyt-c/Cu-BDC composite is far superior than the till date reported electrochemical sensors for NADH. These features qualify Cyt-c/Cu-BDC composite as a promising electrode material for the design of point-of-care NADH sensing devices for clinical diagnostics.</p> Graphical Abstract <p></p>