The rejection of arsenopyrite during chalcopyrite flotation is crucial to prevent the arsenic pollution to the ecosystem. Herein, surface modification by calcium ions coupled with the carboxymethyl cellulose (CMC) were successfully used to achieve Cu-As separation. Flotation results confirmed that the depressive effect of CMC alone towards arsenopyrite was unsatisfactory, but it was greatly enhanced in the presence of calcium ions (Ca²⁺), thus the As content in copper concentrate was lowered significantly. Mechanism research indicated that the wettability of arsenopyrite caused from CMC was strongly enhanced by the pretreatment with Ca²⁺, while that of chalcopyrite increased slightly. Reagent adsorption and FTIR spectra measurements further illustrated the adsorption of CMC on arsenopyrite was facilized by Ca²⁺, but that on chalcopyrite remained very weak. More detailed adsorption mechanisms uncovered by XPS and DFT calculations demonstrated that the Fe, As and S atoms of arsenopyrite were unactive and thus unable support the chemisorption of CMC. Nevertheless, hydroxyl calcium ions adhered on the surface of arsenopyrite after interaction with Ca²⁺. Furthermore, the electrophilic strength of the adhered hydroxyl calcium ions was stronger than that of pristine arsenopyrite surface, ultimately, CMC chemisorbed on arsenopyrite surface through the bridging effect of adhered hydroxyl calcium ions.