Nanoparticle-Mediated Visualization of Endo-Lysosomal Remodeling in Bacterially-Infected Cells

Take a chambered slide containing human epithelial cells with fluorescently labeled endosomes.

Introduce intracellular pathogenic bacteria and incubate, allowing them to adhere to the cells.

The bacteria inject proteins that reorganize the host cytoskeleton, forming membrane protrusions that internalize the bacteria into vacuoles.

These vacuoles extend tubular projections connecting to the endo-lysosomal system, a network of vesicles and organelles regulating intracellular trafficking.

These projections redirect the vesicle transport and remodel the endo-lysosomal system.

Wash to remove extracellular bacteria, then incubate with an antibiotic-containing medium to eliminate any remaining bacteria.

Replace with a serum-free imaging medium containing the antibiotic, and add nanoparticles labeled with a contrasting fluorophore.

Incubate to allow nanoparticle entry via endocytosis and accumulation in the remodeled vesicles.

Wash to remove non-internalized nanoparticles, then incubate in a serum-supplemented imaging medium.

The fluorescent nanoparticles and labeled endosomes enable visualization of the bacteria-induced endo-lysosomal remodeling.

To begin this procedure, measure the OD 600 of the s