Not all tumor vessels are equal. Tumor-associated vasculature includes immature vessels, regressing vessels, transport vessels undergoing arteriogenesis and peritumor vessels influenced by tumor growth factors. Current techniques for analyzing tumor blood flow do not discriminate between vessel subtypes and only measure average changes from a population of dissimilar vessels. We have developed methodologies for simultaneously quantifying blood flow (velocity, flux, hematocrit and shear rate) in extended networks at single capillary resolution in vivo. Our approach relies on deconvolution of signals produced by labeled red blood cells as they move relative to the scanning laser of a confocal or multiphoton microscope and provides fully-resolved three-dimensional flow profiles within vessel networks. Using this methodology, we show that blood velocity profiles are asymmetric near intussusceptive tissue structures in tumors in mice. Furthermore, we show that subpopulations of vessels, classified by functional parameters, exist in, around a tumor and in normal brain.

Key Points AT1413 is a monoclonal antibody isolated from a cured patient with AML that recognizes CD43s, a novel epitope expressed by AML and MDS blasts. AT1413 eliminates CD43s-expressing leukemic blasts in vitro and in vivo and may have potential as a therapeutic antibody.

Current methods to determine cellular cytotoxicity in vitro are hampered by background signals that are caused by auto-fluorescent target and effector cells and by non-specific cell death. We combined and adjusted existing cell viability assays to develop a method that allows for highly reproducible, accurate, single cell analysis by high throughput FACS, in which non-specific cell death is corrected for. In this assay the number of living, calcein AM labeled cells that are green fluorescent are quantified by adding a fixed number of unlabeled calibration beads to the analysis. Using this modified FACS calcein AM retention method, we found EC50 values to be highly reproducible and considerably lower compared to EC50 values obtained by conventional assays, displaying the high sensitivity of this assay.