Overexpression of Cat D in BE-2 cells. (A) BE-2 neuroblastoma cells were either transfected with GFP (control) or with Cat D-GFP and lysosomes stained with lysotracker red. The GFP expressing BE-2 cells (left panels (LysT-Red, GFP-merge), bar = 5 μm) show few lysosomes localized inside the cell body (see Lysotracker red (LysT-red) panels); whereas, the Cat D-GFP transfected cells (right panels (LysT-Red, CD-merge), bar = 5 μm) show build up of lysosomes (see Lysotracker red (LysT-red) panels). GFP-merge and CD-Merge represent the corresponding merged images with green channel. (B) Quantification of lysosomes was performed on BE-2 cells transfected with either GFP or Cat D (n = 10 cells for each group). Unpaired t-test, p < 0.001 (C) Western blot showing the expression of Cat D (~30 kDa, the mature Cat D heavy chain) in non-transfected (control) and transfected (Cat D) BE-2 cells. β-actin was used as loading control. Lower bar graph is representative of 3 individual experiments. Mean and standard error of the mean shown, significance is indicated by ***p < 0.001.