Thiols play critical roles in biological processes. Here, a facile hydrothermal synthesis approach was used for fabrication of flourescent Cu-carbon dots (Cu-CDs), to be employed as accurate thiol Nanosensors. The obtained Cu-CDs were characterized using transmission electron microscopy, UV–visible, photoluminescence and X-ray photoelectron spectroscopy scannings. The highest-occupied- and lowest-unoccupied- molecular-orbitals and the pertaining energy band gap were achieved using analytical chemistry density functional theory. Electrochemical performance of Cu-CDs showed the capability of the fabricred nanoparticles for accurate, sensitive, and selective nanosensing of thiols, as both electrochemical and optical detection tools. The electrochemical peak of Cu-CDs were selectively, rationally and intensively suppressed by thiols, due to the strong electrostatic interaction and covalent bonding between the NH2 functional groups of the Cu-CDs and -SH groups of thiols. On the other hand, the blue photoluminescence emission of Cu-CDs rationally increased, due to detachment of Cu ions from the edges of Cu-CDs in the existance of thiols. According to the above description, the limit of detections (LODs) of 1 nM was achieved, introducing such-prepared Cu-CDs nanosensors, as simple, fast, low-cost, sensitive and selective tools for thiol detection.