High-Resolution Detection of Chlamydia Proteins by Expansion Microscopy

The pathogenic bacterium Chlamydia replicates via an unusual developmental cycle that takes place within a membrane-bound inclusion inside an infected host cell. Immunofluorescence microscopy has revealed many details about the two specialized developmental forms and the recruitment of host organelles to the inclusion, but lacks the high resolution of electron microscopy (EM). In this chapter, we describe the use of an innovative microscopy method, called Tenfold Robust Expansion (TREx) microscopy, to visualize proteins in Chlamydia-infected cells at nanoscale resolution. We present a detailed protocol describing the steps of this expansion microscopy method, including immunostaining, anchoring, gelation, homogenization, expansion, and visualization with a standard confocal microscope.