5–20 wt% trimethoxysilylpropyl octadecyldimethyl ammonium chloride (QAS) was used to modify Poly (ε-caprolactone) (PCL)-gelatin hybrid to fabricate non-leaching antibacterial nanofiber membranes (PG-Q) by electrospinning. The results from scanning electron microscopy (SEM) and transmission electron microscopy (TEM) indicated that the QAS leaded to phase separation between the QAS and PCL. Hydrophilic test demonstrated that the PG-Q nanofiber membranes had hydrophobic surface, which was help for peeling off the dressing from the wound. Additionally, the physical and chemical cross-linking between the QAS/PCL and QAS/gelatin were confirmed by Fourier transform infrared (FTIR), which were good for long lasting antibacterial effect. The PG-Q membranes also showed excellent cell-biocompatibility. Furthermore, compared with pure PCL nanofiber membrane, the PG-Q nanofiber membranes, especially PG-Q15 (QAS: 15 wt%) and PG-Q20 (QAS: 20 wt%), showed a considerable increase in the bacteriostatic rate of S. aureus and P. aeruginosa (more than 99% after 12 h). Therefore, electrospinning non-leaching antibacterial nanofiber membranes could be an optimal choice for antibacterial wound dressing.