The abnormal lattice expansion of commercial polypropylene (PP)/polyethylene (PE)/polypropylene (PP) separator in lithium-ion battery under different charging current densities was observed by in-situ X-ray diffraction. Significant lattice changes of both PP and PE were found during the low current density charging. The capacity fading and the resistance value of the cell measured at 0.025 C (5th retention, 92%) is unexpectedly larger than that at 1.0 C (5th retention, 97.3%) from the electrochemical impedance spectroscopic data. High-resolution scanning electron microscopy is employed to witness the pore changes of the trilayered membrane. Density functional theory calculations were used to investigate the mechanism responsible for the irregular results. The calculations revealed that the insertion of Li-ion and EC molecule into PP or PE are thermodynamically favourable process which might explain the anomalous significant lattice expansion during the low current density charging. Therefore, designing a new separator material with a more compact crystalline structure or surface modification to reduce the Li insertion during the battery operation is desirable.