Peripheral nerve injuries (PNI) are a large-scale problem that affects over one million people around the world. For PNI that require surgical intervention and in the case of long gap injuries, autologous nerve grafts (ANG) have been considered the gold standard for decades. However, the random efficiency of ANG is associated to several drawbacks, for instance, donor site morbidity, limited availability and nerve mismatches, leading to merely 50% rates of success. Keratin, our selected protein, is a highly available fibrous protein found in hair, wool and feathers and it has been used as a potential scaffold material for tissue engineering procedures since it possess cell binding motifs capable of supporting cellular attachment. In this study, an innovative combination of chitosan and keratin (human air) is obtained in order to create novel materials aimed at finding applications in the treatment of PNI. Chitosan/keratin membranes were produced by solvent casting technique and were physicochemical and biologically characterized. Keratin/chitosan membranes topography showed a rough surface, with ridges and pores. FTIR revealed characteristic peaks of keratin and chitosan in the membranes suggesting the presence of both biomaterials in the blend solution. Regarding mechanical properties, the membranes showed mainly elastic behavior and a low ability to dissipate energy, with a considerably high stiffness. In a preliminary screening using L929 cells in both materials, it was observed that cell viability increased with culturing time in chitosan/keratin membranes. In a more specific study including several cell types relevant to peripheral nerve regeneration, such as human Schwann cells (SC), microvascular endothelial cell line (HPMEC-ST1) and human dermal fibroblasts (BJ), there is a significantly higher cell adhesion and metabolic activity after 14 days in chitosan membranes containing 1% keratin, comparing to 100% chitosan membranes. This study demonstrated that keratin-containing biomaterials possess interesting properties for application in Peripheral Nerve Regeneration.