Summary: Copper-loaded carboxymethyl-chitosan (CMCS-Cu) nanoparticles were successfully prepared by chelation under aqueous conditions. The effect of degree of deacetylation and substitution, the molecular weight of CMCS, CMCS concentration, Cu(II) ions concentration, pH value of the solution, as well as temperature, on the morphology of the yielded particles were systematically investigated. The physicochemical properties of the particles were determined by size and zeta potential analysis, FTIR analysis, DLS, TEM, SEM and XRD pattern. FTIR and XRD revealed that Cu (II) ions and CMCS formed a chelate complex. The size of CMCS-Cu particles shows a good consistency by DLS, TEM, and SEM. The nanoparticles with the size of about 70 nm have been prepared at 0.13 wt% CMCS, 16 mmol/L Cu(II) ions, pH value 4.56 at 25 °C. The antibacterial activity of CMCS, CMCS-Cu normal particles with the size of about 1000 nm and CMCS-Cu nanoparticles with the size of less than 100 nm against Staphylococcus aureus was evaluated by vibration method. Results show that the antibacterial efficiency of nanoparticles reached 99%, which is much more efficient than 68.9% of the normal one and 6.1% of CMCS. CMCS-Cu nanoparticles were proved to be a good novel antibacterial material.