Casein fibers, produced from plant or animal protein, have some good features such as comfort , high moisture traort, lightweight (density = 1.3 g/cm 3 ), good for health (pH near to skin), tiny insects and fungus resistant (increase the longevity of the fabric), high thermal insulation, anti-bacterial, and so on. Casein has strong molecular structure due to the hydrogen bonds, so it could not be purely electrospun with common solvent. Therefore, to overcome this problem, the link between molecular and inter-molecular should be reduced. This purpose can be achieved by adding the polymer that structurally is non-homologous to the structure of protein. For this purpose, Xie and Hsieh produced casein nanofibers by adding PVA and PEO polymers as auxiliary material for electrospinning. In this work, we produced electrospun nanofibers yarn from casein which is not possible with conventional spinning systems. Polyethylene Oxide (PEO) and Polyvinyl alcohol (PVA) were used with the Casein as auxiliary polymer to allow spinning of Casein. Optimal conditions including process parameters such as voltage, feed rate, distance between two nozzles and solution parameters including concentration and blending ratio was optimized to produce good quality yarn. Morphological structure of PEO/Casein and PVA/Casein with various blending ratio with different PEO, PVA and Casein concentration was investigated. The morphology of different nanofiber web and nanofiber yarn was observed using SEM and fiber diameter was measured by Digimizer software. Results showed that PEO (26 wt %) / Casein (10 wt %) 10:90 was the best composition that produced yarn with smooth structure and minimum bead in its structure. This sample chosen as the desired sample. In order to improve the properties of the desired yarn, the process of crosslinking was done. To achieve this goal, this yarn crosslinking with two crosslinker, Formaldehyde and Diisocyanate. Diisocyanate was used in two concentration, 20% and 40%. Then mechanical properties of yarn such as stress, strain, work of rapture, elastic modulus were studied on all yarn samples, before and after of crosslinking. At last the best crosslinker is Diisocyanate with the concentration of 40% was chosen as best crosslinker for crosslinking of desired yarn.Finally fracture morphology of nanofiber yarn has investigated. Taken images were used to analyze the broken ends of the yarns and showed that the desired sample nanofiber yarn with Diisocanate at 40% concentration has the brittle structure in compared with desired sample nanofiber yarn without crosslinker. Key words : Casein Fibers, Electrospinning, PEO, Crosslinking, Facture morphology