EGFRvIII increases the microvessel density and vascular permeability in tumors. (A) Frozen sections of tumor tissues extracted from the mice implanted with LN229 cells transfected with mock (a), wtEGFR (b), EGFRvIII (c) were prepared and immunohistochemically stained for CD31, an endothelial cell marker. Scale bars show 100 μm. (B) Quantitative analysis of the microvessel density in the tumors was performed. At least three fields were selected from each section. Vessel area/mm2 was measured as the average of the values in four sections/mouse. Data shown are the means ± SEM (n = 5 mice). Significant differences are shown: *** p<0.001. (C) Permeability of the tumor blood vessels in LN229-tumor bearing mice. The mice were intravenously injected with TexasRed conjugated dextran (Mw. 70,000, 25 mg/mouse, red, d-f). Subsequently, 6 h after the dextran injection, Alexa647-conjugated isolectin IB4 (green, a-c), a probe used for specific labeling of endothelial cells, was administered. The mice were sacrificed, and frozen sections were prepared and observed by fluorescent microscopy. The cell nucleus were labeled with DAPI (blue, g-i). The merged images are shown (j-l). Scale bars show 50 μm.