A separator film for high-performance Li-ion batteries was prepared by electrospinning. The film had a hybrid morphology of silica nanofibers (SNFs) and alumina nanoparticles (ANPs), with a smooth surface, polymer-free composition, high porosity (79%), high electrolyte uptake (876%), and excellent thermal stability. Contact angle measurements demonstrated the better immersion capability of the SNF-ANP separator film for commercial liquid electrolytes than a commercial CELGARD 2500 separator film. Moreover, compared to the commercial CELGARD 2500 separator, the ionic conductivity of the SNF-ANP separator film was nearly three times higher, the bulk resistance was lower at elevated temperature (120 °C), the interfacial resistance with lithium metal was lower, and the electrochemical window was wider. Full cells were fabricated to determine the cell performance at room temperature. The specific capacity of the full cell with the SNF-ANP separator film was 165 mAh g−1; the cell was stable for 100 charge/discharge cycles and exhibited a capacity retention of 99.9%. Notably, the electrospun SNF-ANP separator film can be safely used in Li-ion or Li-S rechargeable batteries.