Comparative imaging of a bacterial surface‐located GFP fusion protein by epifluorescence and scanning near‐field optical microscopy

Summary IcsA is an autotransporter protein that plays a role in the virulence of Shigella bacteria. We have examined the cellular localization of a fusion of an IcsA fragment to the green fluorescent protein (GFP) expressed in Escherichia coli using a dual epifluorescence and scanning near‐field optical microscope. By combining the data obtained from far‐field with near‐field microscopy of the same sample, discrimination between surface‐bound fusion proteins and fusion proteins located in the cellular cytoplasm becomes possible. Furthermore, and for the first time, the inherent advantages in resolution of the near‐field images provides highly specific details of the location of a GFP fusion protein on a bacterial cell surface.