... MedicalScience, University of Tokyo, Japan) for providingpCAG-Cre; Dr Ikuo Yana (Institute of MedicalScience, University of Tokyo, Japan) for help in per-forming endothelial tube formation experiments. ... Tokyo,Japan). For immunocytochemistry, cells were fixed on ice with4% PFA in NaCl ⁄ Pi for 10 min, incubated in methanol at)20 °C for 20 min, and rehydrated in NaCl ⁄ Pi. For detec-tion of Prox-1, cells ... primary cells. Thus, ‘immortalization’ techniques have been stronglyrecommended for most analyses that require a largequantity of transcripts, proteins or cells. For immortal-ization of ECs,...