Visualization of Peptidoglycan Structures of Escherichia coli by Quick-Freeze Deep-Etch Electron Microscopy

Peptidoglycan (PG) is an essential component of the bacterial cell wall that protects the cell from turgor pressure and maintains its shape. In diderm (gram-negative) bacteria, such as Escherichia coli, the PG layer is flexible with a thickness of a 2-6 nm, and its visualization is difficult due to the presence of the outer membrane. The quick-freeze deep-etch replica method has been widely used for the visualization of flexible structures in cell interior, such as cell organelles and membrane components. In this technique, a platinum replica on the surface of a specimen fixed by freezing is observed using a transmission electron microscope. In this chapter, we describe the application of this method for visualizing the E. coli PG layer. We expect that these methods will be useful for the visualization of the PG layer in diverse bacterial species.