In the pre-formulation stages of cosmetic products, in vitro permeability tests correspond to an important alternative for making better decisions regarding the stability, performance and biocompatibility of these products. Moreover, these tests allow us to establish whether a cosmetic ingredient can penetrate the different layers of the skin, which is essential in these products. This study was focused on the extraction, characterization and preservation of rabbit ear membranes and their subsequent performance comparison against two synthetic membranes (cellulose and Strat-MTM). For this, the rabbit ear stratum corneum was isolated and characterized histologically, using the Hematoxylin and Eosin (HE) staining protocol along with light microscopy and image analysis. Then, the Frank diffusion cell (FC) model was employed to evaluate and compare the permeability of the model compound Naproxen sodium (NPX) between the three membrane systems. The results show that NPX permeability is strongly affected by the type of membrane, and the implementation of rabbit ear membranes shows high reproducibility, demonstrating that this model could be implemented during pre-formulation studies of cosmetic products.