Fabricating metal-chelating affinity materials with excellent protein adsorption and isolation properties is of great significance in numerous biotechnological fields, yet still face huge challenges. Herein, we successfully developed a new kind of nanofiber based metal-chelating affinity membranes by orderly modifying the electrospun regenerated cellulose nanofibrous membranes (RC NFMs) with intermediate bridging agent of cyanuric chloride (CC), chelating agent of iminodiacetic acid (IDA) and affinity ligand of Fe3+ ions. The resultant Fe3+ ions immobilized RC NFMs (RC-CC-IDA-Fe3+ NFMs) exhibited a high lysozyme adsorption amount of 365 mg g(-1) within the fast equilibrium adsorption time of 4 hours. Meanwhile, the performance of RC-CC-IDA-Fe3+ NFMs could be facilely regulated by controlling the properties and concentration of protein solutions. Moreover, the obtained nanofibrous affinity materials also possessed superior cycle performance and excellent regeneration ability. Undoubtedly, the successful fabrication of efficient RC-CC-IDA-Fe3+ NFMs might provide a new kind of affinity adsorbent for protein purification and other potential biotechnological fields.