The surface property regulation of fluorite and calcite minerals is an important method to increase their flotation performance differential for potentially solving the conundrum of their flotation separation. Therefore, the flotation separation of fluorite and calcite by using various metallic ions including Cu2+, Al3+, Pb2+, Fe2+ and Fe3+ as surface regulation agents was systematically investigated in this research. Based on the results of quantitative mono-mineral micro-flotation experiments, a great gap value of 58.12 % between the flotation recovery of fluorite and calcite could be obtained simply by introducing Fe2+ (300 mg/L) pretreatment with NaOL (80 mg/L) collector and no extra depressants used at slurry pH = 6, demonstrating the effectiveness of Fe2+ pretreatment for enhancing the flotation separation efficiency. Furthermore, a wide array of analytical and detection techniques were employed to uncover the metallic ions differential regulation mechanism. The obtained findings indicated that the fluorite surface was relatively clean and smooth without obvious surface corrosion by metallic ions. However, under the combined action of OH- and CO32- as a result of the surface dissolution of calcite and O2 in the air, those metallic ions can precipitate as Cu4SO4(OH)6 & sdot;2H2O, PbCO3, Al (OH)3, and Fe(OH)3 on the surface of calcite through the chemical reaction, thereby shielding the original flotation active sites (calcium atoms) of the calcite surface and affecting its flotation behavior in the sodium oleate system. In particular, the metallic compound attached on calcite surface after Fe2+ or Fe3+ pretreatment was more evenly and tightly than that after Cu2+, Pb2+, and Al3+ pretreatment, thus forming a relatively dense surface passivation layer to significantly hinder the subsequent adsorption of oleate anion (OL-) on calcite surface, resulting in greater difference in the flotation performance of two minerals and consequent higher separation efficiency. This research provided new insights into the differential surface regulation of calcite and fluorite by various metallic ions, which can be potentially applicable to the flotation separation of fluorite and calcite.