Many industries use copper metal ions (Cu2+ ions), and their salts are utilized as supplemental materials in both agriculture and medicine. Identifying and monitoring these Cu2+ ions in biological and environmental specimens is crucial due to their association with several health issues. In this investigation, we have designed a simple quinoline‐based receptor (E)‐3‐(((2,4‐di‐tert‐butyl‐5‐hydroxyphenyl)imino)methyl)‐6‐methoxyquinolin‐2(1H)‐one (QAP) containing imine functional groups to inspect its capability to identify metal ions in a semi‐aqueous medium. The photophysical characteristics and structural confirmation of the receptor QAP were investigated using various spectroscopic techniques. Among various metal ions, the receptor QAP displayed an intense color shift from slightly yellow to strong yellow in the existence of Cu2+ ions, as visualized by the nude eye. Furthermore, the fluorescence spectral maximum wavelength at 485 nm and the strong cyan fluorescence color were quenched upon introducing Cu2+ ions. The alteration in the spectral and colorimetric features of QAP with Cu2+ ions is due to coordination complex formation. The present sensor shows the linear range from 3 to 69 μM, subsequent in a computed limit of detection as 3.16 nM, which is much lower than that of the maximum threshold of Cu2+ ions in drinking water set by WHO. Therefore, the receptor can respond to Cu2+ ions sensing in two ways: by changing color and by quenching fluorescence. The binding mode of the Cu2+ ions to the functional groups of the receptor QAP is a 1:1 stoichiometry, according to ESI‐mass, Job's plot analysis, and density functional theory (DFT) computations. The practical utility of the fluorescent receptor QAP was applied for Cu2+ ions determination in environmental samples (drinking, tap, and dam water) and cancer cells (HeLa cells).