Matrigel Plug Angiogenesis Assay in vivo

BALB/c nu/nu mice were administered a subcutaneous injection of 400 μL mixture containing 2 × 10⁶ EA.hy 926 cells premixed with a Corning Matrigel Matrix phenol red-free (Cat. No. 356237), IL-8 (100 ng/ml), and UDCA (50 μM). Nude mice were randomly divided into three groups (the Control group, IL-8 group, and IL-8+UDCA group). Matrigel plugs were removed and imaged after 10 days. A Bestbio reagent kit (Bestbio, CHA) was applied to detect the hemoglobin content of the matrigel plugs according to the manufacturer’s protocol. Plugs were also snap frozen in the presence of optimum cutting temperature medium before sectioning and stained by hematoxylin and eosin (HE). Immunohistochemistry (IHC) assay was also performed to determine the expression of CD31, VEGF, and vWF (Santa Cruz Biotechnology, CA). Guide for the Care and Use of Laboratory Animals of the National Institutes of Health was strictly complying with in the in vivo experiment which was approved by the Committee on the Ethics of Animal Experiments of the Second Military Medical University.