Co-localization of LAMP-2 and LipidTOX. (A-D) Control ARPE-19. (E-H) 20 μg/ml chloroquine-treated ARPE-19. Control phase contrast (A) shows no vacuolation and LAMP-2 (B, green), and LipidTOX(C, red) staining is observed throughout cell. D is fluorescent overlay. Chloroquine-treated cell in phase contrast (E) shows intense vacuolation (white arrows). These vacuoles co-localize with LAMP-2 staining (F, corresponding white arrows). Chloroquine treatment also induces dense body formation (E, yellow arrows). Dense formations co-localize with neutral lipid (G, corresponding yellow arrows). Overlay (H) shows no overlap between lipids and vacuoles. Bar size = 25 microns.