A planar electrochemical sensor, based on flower-like CuO nanostructures growth “in situ” on a commercial screen printed carbon electrode, was fabricated by an easy and effective technique and employed for the non-enzymatic determination of glucose. The prepared CuO nanostructures were characterized by scanning electron microscopy (SEM) and X-ray diffraction (XRD). The carbon electrode modification with CuO was optimized by investigating the effect of the number of deposition cycles of precursor and their concentration. The electrodes modified by in situ growth of CuO were compared to an electrode prepared by simple deposition of CuO powder previously synthesized by the same technique. Cyclic voltammetric and chronoamperometric tests demonstrated that the in situ growth of CuO leads to excellent electrochemical performance toward glucose oxidation in 0.1 M KOH solution. The best sensor, if operated at an applied potential of 0.6 V, has a sensitivity of 1460 μA·mM −1 ·cm −2 and a 2.5 μM detection limit (at an S/N ratio of 3). Tests carried out within six months revealed an excellent long-term stability. This suggests that the method applied to modify the carbon electrode represents a useful tool for fabrication of an inexpensive and reliable non-enzymatic glucose sensor. Graphical abstract Schematic of a glucose sensor fabricated by growing flower-like CuO nanostructures in-situ on a screen printed carbon electrode (SPCE). The sensor showed excellent sensitivity and long-term stability.