Amphiphilic copolymer Pluronic F127 was used as multifunctional additive to fabricate antifouling polyethersulfone (PES) ultrafiltration membranes. The scanning electron microscopy (SEM) photographs showed an asymmetric morphology of PES/Pluronic F127 membranes. The static water contact angle measurements and X-ray photoelectron spectroscopy (XPS) analysis confirmed surface hydrophilic modification of the membranes. The protein rejection experiments indicated that the pore sizes of the skin layer were enlarged with the increase of Pluronic F127 content. The dual role of Pluronic F127 as surface modifier and pore-forming agent was distinctly observed and tentatively analyzed. During the dissolution and subsequent coagulation process, some Pluronic F127 molecules are bound to and/or tangled with PES chains due to the presence of hydrophobic PPO blocks, and then spontaneously segregate to the membrane/water interfaces due to the presence of hydrophilic PEO blocks. Meanwhile, the majority of Pluronic F127 molecules self-assemble into spherical micelles with core/shell configuration. These micelles are extracted into the coagulation bath during the coagulation process and the spaces they once occupied will turn into the pores of the membranes. In summary, a facile method for fabricating a series of PES/Pluronic F127 membranes with controlled flux and excellent antifouling property was developed.