Scanning near field optical microscopy (SNOM) has been used to visualize phase-separated regions in a monolayer protein film. The film was created at an air-water interface during displacement of the protein bovine serum albumin (BSA) by the nonionic surfactant Tween 20. Contrast was generated in the SNOM images By fluorescent labeling of the protein. The validity of the SNOM methodology has been confirmed by a qualitative and quantitative comparison of the displacement process as visualized by SNOM and atomic force microscopy (AFM) on fluorescently labeled BSA displaced by Tween 20 and AFM data on native BSA displaced by Tween 20. The data suggest that SNOM studies of labeled proteins can be used to investigate the behavior of particular protein species in complex multicomponent interfacial films.