2.13 Histological and immunofluorescence analysis
The vascular structures were fixed with 4% v/v paraformaldehyde after culture for 7 days and cut into 15μm section with a cryostat (CM1950, Leica, Germany) at -20℃. Slices were stained with hematoxylin and eosin staining (H&E) and Masson’s trichrome staining. Image were collected by light microscopy (DM4000b, Leica, Germany).
For immunofluorescence staining, HUVECs were labeled with mouse anti-human CD31 (1:50, Abcam, USA) and SMCs were labeled with rabbit anti-human α-SMA (1:200, Abcam, USA). Alexa Fluor 488 goat anti-mouse (1:500, Abcam, USA) and Alexa Fluor 594 Donkey anti-rabbit (1:500, Abcam, USA) were used as secondary antibodies. All the section were counterstained with DAPI. Fluorescence images were obtained with a fluorescent microscope (Leica, DM8000, Germany). The sections without primary antibody staining were used as control.