The antimicrobial polypeptide epsilon-poly(L-lysine) (epsilon-PL) was electrostatically incorporated to poly(acrylic acid) (PAA)/poly(vinyl alcohol) (PVA) electrospun nanofibers. epsilon-PL loading and distribution was assessed by infrared spectra, zeta-potential measurements and the primary amino reactive dye fluorescamine. Functionalized fibers with 485 +/- 140 nm diameter, could be loaded with 0.57-0.74 g epsilon-PL (g dressing)(-1) that released at a constant rate of 5.4 +/- 2.8 mg epsilon-PL (g dressing day)(-1). Such a dressings resulted in two orders of magnitude lower bacterial colonization than non-functionalized PAA-PVA after 14 days of incubation. Bacterial impairment was attributed to the damage of cell membranes and the formation of intracellular reactive oxygen species. epsilon-PL functionalized nanofibers did not display cytotoxicity to human corneal epithelial cells, HCEpC, in 24 h MTT assays. However, the viability of rapidly growing tumoral HeLa cells decreased > 50% under the same conditions. The prepared biocompatible nanofibrous dressings with durable antibacterial activity show potential application as wound dressings and other biomedical uses.