There is a consistent demand for developing highly sensitive, stable, cost-effective, and easy-to-fabricate creatinine sensors as creatinine is a reliable indicator of kidney and muscle-related disorders. Herein, we reported a highly sensitive and selective non-enzymatic electrochemical creatinine sensor via modifying poly(3,4 ethylenedioxythiophene) polystyrene sulfonate (PEDOT:PSS) coated indium tin oxide (ITO) substrate by zeolitic imidazolate framework-8 nanoparticles (ZIF-8 NPs). The topography, crystallinity, and composition of the sensing electrode were characterized by field emission scanning electron microscopy (FESEM), atomic force microscopy (AFM), X-ray diffraction (XRD), and X-ray photoelectron spectroscopy (XPS). The peroxidase-like activity of ZIF-8 nanoparticles enabled it to detect creatinine forming a zinc-creatinine composite. The electrochemical behavior and sensing performance were evaluated by amperometric and impedimetric analysis. The sensor obtained a sufficiently low limit of detection (LOD) of 30 µM in a clinically acceptable linear range (0.05 mM–2.5 mM). The interference study demonstrated high selectivity of the sensor for creatinine concerning other similar biomolecules. The sensing performance of the creatinine sensor was verified in the actual human serum, which showed excellent recovery rates. Hence, the magnificent performance of ZIF-8 based non-enzymatic creatinine sensor validated it as a responsible entity for other complicated renal markers detection.