Figure 2

The combination of PS-341 and Doxil^® induces enhanced apoptosis in vivo. The impact of vehicle, PS-341 alone, Doxil^® alone, or the combination, was studied in a murine xenograft model of human breast cancer established using BT-474 breast carcinoma cells. Apoptosis was evaluated in tumor sections 24 hours after the indicated treatments by detection of single stranded DNA fragmentation using the murine monoclonal antibody Mab 3299 [45] (Chemicon International, Temecula, CA, USA). Single stranded DNA associated with programmed cell death (red) is shown, along with total nuclear DNA (blue), the latter detected using 4,6-diamidino-2-phenylindole (Vector Laboratories, Burlingame, CA, USA). Slides were visualized using an ultraviolet Zeiss Axioplan fluorescence microscope (Carl Zeiss Optical, Inc., Chester, VA, USA). Separate photographs were taken with appropriate filters for blue nuclear staining and red single-stranded-DNA staining, overlayed using Adobe Photoshop software, and displayed as a fusion image at 10 × magnification.