Anti ve cadherin af647 antibody clone bv13

25 µg of anti-VE-cadherin-AF647 antibody (clone BV13, Biolegend) were injected retro-orbitally in 8–10-week-old male C57BL/6J (CD45.2⁺) mice under anaesthesia 8 min before they were killed and the organs harvested. For flow cytometry and cell sorting, organs, including lungs, brain, kidney and liver were minced and incubated with collagenase-A (25 mg ml⁻¹), Dispase II (25 mg ml⁻¹), and DNase (250 µg ml⁻¹) (Roche) at 37 °C for 20–30 min to create a single-cell suspension. Cells were filtered through a 40-µm filter immediately before counter staining. Single-cell suspension was first blocked with an antibody against CD16/32 (2.4G2) before antibody staining with anti-mouse CD31-PE-Cy7 (390, Biolegend), anti-mouse TER119 (TER119) and anti-mouse CD45 (30-F11). Haematopoietic and erythroid cells were removed via CD45 and TER119 gate exclusion, and adult mouse endothelial cells were defined and sorted as VEcad⁺CD31⁺CD45⁻TER119⁻ cells. Resulting adult mEC cultures were cultured on fibronectin-coated (Sigma-Aldrich) plates in mEC media. Purity of mECs and absence of contaminating haematopoietic cells were confirmed by flow cytometry and confocal microscopy.