MXD3 protein expression was evaluated by immunochemistry as previously described (10 (link)). Briefly, cells were fixed with 10% buffered formalin, smeared onto slides, and blocked with 10% FBS. Slides were incubated with anti-MXD3 monoclonal mouse antibody (Antibodies Inc, Davis, CA) overnight at 4 °C. Secondary goat anti-mouse antibody conjugated to Alexa Fluor 488 (A488) (Thermo Fisher Scientific) incubation was performed at room temperature for 3 hours. MXD3 protein expression was quantified by measuring the fluorescent image intensity of all cells within a representative image (TIFF) and averaged to obtain the mean fluorescence intensity. Measurement and quantification was performed using ImageJ (NIH) (23 (link)). Briefly, individual cell boundaries were marked and the mean fluorescence intensity (MFI) was measured for each cell. A background signal (without cells) was subtracted from each MFI. Corrected MFI for each cell was then averaged per experiment and treatment type.
Tumor microarrays with 18 human neuroblastoma tissue samples were stained for MXD3 at the University of California, Davis (UC Davis) Cancer Center Pathology Core.
Tumor microarrays with 18 human neuroblastoma tissue samples were stained for MXD3 at the University of California, Davis (UC Davis) Cancer Center Pathology Core.
