This work shows that electrografted polymethacrylonitrile films can be used for direct immobilization of proteins through spontaneous reaction between the nitrile groups from the film and the amino groups from the amino acids of the protein, in alcoholic basic medium. The formation of the resulting imino-ethers groups is followed by XPS measurements and assessed by high precision calculations of core electron binding energies. The final protein film is stable toward aqueous rinsing and can be used for protein-substrate recognition.