The invention relates to an HUVEC (
human umbilical vein endothelial cell) separation, culture and subculture method belonging to the technical field of
biology. The method comprises the following steps: (1) separation of HUVEC: taking a fresh
umbilical cord, cleaning with a sterile PBS (
phosphate buffer solution) containing
penicillin-
streptomycin, adding
collagenase to perform water bath
digestion at 37 DEG C, centrifuging the
digestion solution, then adding an M199 culture medium, transferring into a
gelatin-coated culture
bottle, and culturing in a CO2
incubator at 37 DEG C; (2) culture of HUVEC: after the HUVEC is cultured at 37 DEG C for 24 hours, pouring out the culture medium, cleaning with the PBS, adding a fresh M199 culture medium, and afterwards, changing the culture medium once every two days, wherein the HUVEC can be subcultured generally after being cultured for 5-7 days; and (3) subculture of HUVEC: pouring out the culture medium, cleaning with the PBS, adding the
digestion solution to digest the
cell, adding a DMEM (dulbecco's modified
eagle medium) containing serum to terminate the reaction once the
cell is rounded, centrifuging the digested
cell, and adding a fresh culture medium, wherein the HUVEC subcultured for 2-3 generations is used for experiments. The separated HUVEC is economical and practical, is simple and easy to use, and is beneficial to obtaining required in-vitro
experimental model cells.