Figure 3

Co-localization of fluorescently labeled CPS (FluoSpheres, 25 μg/ml in DMEM + 2%FBS) and lysosomes identified by LysoSensor and cathepsin B activity by CV-(RR) ₂ substrate in EAhy926 cells after 4h of exposure (a) and 24h (b) after particles were removed from the incubation medium. a: Co-labelling of FluoSpheres (FS red) with LysoSensor as marker for lysosomes (green). Scale bar: 10 μm. After 4h co-localization (yellow) is seen for 20 nm and 40 nm FS while 100 nm, 200 nm and 500 nm FS are seen predominantly at the cell periphery. b: After 24h more 100 nm-500 nm FS co-localize with lysosomes and only 500 nm FS were seen at the cell periphery. c: quantification of the co-localization by Metamorph® software: the pH-dependent dye LysoSensor (Sensor, green) and enzymatic activity for the lysosomal enzyme cathepsin B (CatB, red) are used as markers for lysosomes. Co-localization rates of FS with LysoSensor were slightly higher than those with CatB substrate. After 4h significant differences in the co-localization rates were seen in the combinations: 20 nm FS versus 200 nm and 500 nm FS and 40 nm FS versus 100nm, 200 nm and 500 nm FS. After 24h only the differences between 20 nm and 40 nm FS on the one hand and 500 nm FS were significant. Particles with significant differences in the co-localization rates (p < 0.05) are linked by brackets.