This work describes Ni–Ce–Cu metallic–organic framework (MOF) for the detection of non-essential amino acid l-cysteine. The tri-metallic Ni–Ce–Cu MOF was synthesized via a solvothermal method. The cyclic voltammetry and the differential pulse voltammetry techniques were used to examine the electrochemical detection of l-cysteine. The Ni–Ce–Cu MOF shows an oxidation peak in PB solution at pH 3.0 between the potential range of 0.0 and 0.7 V and strong electro-catalytic activity toward the oxidation of l-cysteine across a wide linear range of 0.1 to 250 nM and low detection limit (LOD) was calculated of 1.56 nM. The analysis of l-cysteine in milk and egg yolk samples showed with recovery range of 96.75–103.5% and 97.78–99.43% with RSD% of 2.3–3.2% and 2.7–7.2%, respectively. These results show the Ni–Ce–Cu MOF has high selectivity for l-cysteine detection in milk and egg samples.