Visualization of phospholipid domains in Escherichia coli by using the cardiolipin-specific fluorescent dye 10-N-nonyl acridine orange

Cardiolipin (CL)-specific fluorescent dye 10-N-nonyl-acridine orange (NAO) was used to visualize CL distribution in Escherichia coli cells of different phospholipid compositions. In a filamentous mutant containing only anionic phospholipids, green fluorescent spots were observed along the filaments at approximately regular intervals. Three-dimensional image reconstruction obtained by optical sectioning and a deconvolution algorithm revealed NAO-binding domains in the plane of the cell membrane. Substantial red fluorescence emission of bound NAO supported labeling of CL-containing domains. These structures were not found in mutants deficient in CL biosynthesis. The domains were also observed mostly in the septal region and on the poles in cells of normal size with wild-type phospholipid composition.