A new antimicrobial bilayer film was developed using chitosan, cellulose, and nisin. Chitosan solution containing nisin (500 and 1000 mu g/mL) was prepared by sol-gel method and then the solution was coated on cellulose paper by dip coating method. A chitosan-cellulose film without antimicrobial had no inhibitory effect on Listeria monocytogenes, whereas, the incorporation of nisin made a significant increase (P<0.05) in antimicrobial characteristics of the films. Moreover, no significant differences were shown on antimicrobial activity of developed films during the storage at 4 degrees C for one month. However, the addition of nisin showed a significant increase in the swelling index and solubility of bilayer film. Scanning electron microscope images revealed a uniform coating of chitosan solution on cellulose paper. The FTIR analysis also confirmed successful introducing and binding of the nisin in double layer film. Films with 1000 mu g/mL of nisin completely inactivated the initial (similar to 5 log(10) CFU/g) counts of L. monocytogenes on the surface of Ultra-filter white cheese after storage at 4 degrees C for 14 days. We concluded that nanocomposite film of chitosan-cellulose containing nisin has novel antibacterial activity and can be used for packaging in cheese. (C) 2017 Elsevier B.V. All rights reserved.