Normal large rabbit aortic smooth muscle cell culture

Experimental Materials:

1. Normal large rabbit aorta

2. 1×PBS containing no Ca 2+ and Mg 2+ , adding 200,000 IU/L penicillin, 200 mg/L streptomycin, pH 7.2

3. Digestive juice: 0.125% trypsin, 0.1% collagenase I

4. Cell culture flask (T25)

5. Scalpel, anatomical scissors, anatomical sputum, ophthalmic scissors, ophthalmology é•Š

6. Screen (100 mesh)

7. Centrifuge tube (15ml, 50ml)

experimental method:

1. The rabbits were sacrificed, the aorta was isolated, and washed three times with pre-cooled 1×PBS (pH=7.2) containing double antibody. To wash away the surface blood and mucous membranes;

2. Cut the cleaned aorta into 2.5 ~ 3cm segments and peel off the outer membrane;

3. Place the vascular segment of the adventitia into the culture dish, cut the wall of the tube longitudinally, and then bring the endometrium up, and gently scrape the endometrial cells with a small blade. The membrane was washed 3 times with 1×PBS, and the middle membrane was cut into 1 mm 3 tissue pieces;

4. Move the tissue block into a 50ml centrifuge tube, add an appropriate amount of 0.1% collagenase I solution, and digest it in a 37 ° C incubator for 1 ~ 3 hours until the tissue block becomes flocculent;

5. 1000r/min, centrifuge for 5min, remove the supernatant. Then add 0.125% trypsin solution, stir and digest in a 37 ° C water bath for 5 ~ 10min. The digestion was terminated with a serum-containing medium, and the pipette was gently pipetted for 2-3 minutes to obtain dispersed smooth muscle cells;

6. Pass 100 mesh sieve, collect the filtrate, centrifuge at 1000r/min for 5min, discard the supernatant;

7. Resuspend the cells with the culture medium, adjust the cell concentration to 1×10 5 /ml, inoculate the culture flask, and incubate in a 37 ° C, 5% CO 2 incubator;

8. Change the liquid for the first time after two days, then change the liquid once a day;

9. PriCells-primary smooth muscle cell special basal medium

10. PriCells-Primary Smooth Muscle Cell Culture Special Additives

11. PriCells-Primary Cell Isolation Kit

12. PriCells-Primary Cell Identification Kit