While glucose detection is ruled by enzyme-based glucose test strips, the chemically and thermally sensitive enzymes motivate the researchers to inspect chemically and thermally stable metal oxides as alternate sensing constituents for non-enzymatic glucose detection. Here, we report a synthesis of spinel nickel cobalt oxide (NiCo2O4) nanosheets-based catalyst for enzyme-free glucose sensing application via sonochemical approach using graphene oxide (GO) as nanosheets template and zeolitic imidazolate frameworks (ZIF-67) type nickel-cobalt metal-organic frameworks as oxide precursors (the catalyst named as gt-NiCo2O4 NSs). The physical characterization techniques confirmed that the as-synthesized gt-NiCo2O4 exhibited uniform porous and nanosheets morphology which can be obtained due to the ZIF-67 and GO template, respectively. Cyclic voltammetry and amperometric techniques employed for glucose oxidation at + 0.55 V using gt-NiCo2O4 NSs electrode in 0.1 M KOH. This sensor exhibited an excellent linear range (150 nM–8.86 mM), sensitivity (729.72 µA mM−1 cm−2) and lower detection level (7.35 nM; S/N ratio = 3) to glucose. Interestingly, the gt-NiCo2O4 NSs electrode displayed good recoveries to glucose in human blood serum and urine samples. Hence, the results indicates that gt-NiCo2O4 NSs is a promising electrode matrix with high sensitivity, excellent stability and good reproducibility that can enable the progress of enzyme-free glucose sensors.