Sr flivo

Multiphoton imaging of mammary tumors was done as described previously [56] . MDA-MB-231 cells labeled with Dendra2- or cortactin-GFP were injected into the mammary fat pads of 5–7 week old SCID mice. After 10–12 weeks, we injected Texas Red 70 kDa dextran (250 nmol in 100 µl PBS per injection) [5] (link) for macrophage labeling and performed skin flap surgery 2 h later on anesthetized animals. Exposed mammary tumors were positioned on top of a coverslip on an inverted microscope and imaged continuously using a custom-built two-laser multiphoton microscope for up to 3–5 h. In some experiments, additional tail-vein injections were done for blood vessel labeling using Texas Red 70 kDa dextran (Invitrogen; 250 nmol in 100 µl PBS per injection), MMPSense 680 (Perkin Elmer; 2 nmol in 150 µl PBS per injection), SR-FLIVO (Immunochemistry Technologies; 1∶10 dilution, 2 h prior to imaging), or the pan-MMP inhibitor GM6001 (Milipore; 1 µmol in 100 µl injection). A stock solution of GM6001, 500 mM in DMSO, was diluted in sterile PBS before tail vein injection. Post-surgical injection assures that compounds are only present in tumor blood vessels that are intact, connected to tumor vasculature and flowing at the time of injection.