Experimental Perspectives on Direct Visualization of Endosomal Rupture

Endosomal escape continues to be a limiting factor in the therapeutic use of nanomaterials. Assays to visualize endosomal escape often do not decouple the endosomal/lysosomal disruption from the release of payload into the cytosol. Here, we discuss three approaches to directly probe endosomal/lysosomal rupture: calcein dye dilution, lysosome size quantification and endosome/lysosome membrane integrity visualized with a genetically engineered cell line. We apply the three assays to endosomes/lysosomes ruptured via osmotic pressure and photochemical internalization.