Primary human amniotic cell (HAM) primary cell culture
PriCells: Normal human amnion cells (HAM) primary cell culture Experimental Materials: experimental method: Yellow Wine Without Caramel Colour Yellow Rice Wine,Jia Fan Wine,Shaoxingtian Chun Wine,Yellow Wine Without Caramel Colour ZHEJIANGGUYUELONGSHAN SHAOXING WINE CO.,LTD. , https://www.chinashaoxingwine.com
Human placenta
2. PBSA/GASP: antibiotic-containing PBSA (gentamicin, 50 μg/ml; amphotericin B, 1.25 μg/ml; streptomycin 100 μg/ml; penicillin 100 U/ml);
3. Glycerol;
4. DMEM containing 50% glycerol;
5. Trypsin / EDTA;
6. Millicell microporous membrane tissue culture inserts;
7. Plastic scrapers and sterile cotton swabs;
1. Wash the tissue with PBSA/GASP;
2. Separating the human amniotic membrane and separating the amniotic membrane from the placenta with a gloved hand. The separated thin layer of amniotic membrane includes epithelial cells, basement membrane and some stromal tissue;
3. Use a sterile cotton swab to remove the stromal tissue under the basement membrane so that the separated amniotic membrane is as thin as possible;
4. In the period of monitoring the donor serum to exclude the disease, the crude isolated human amniotic membrane can be stored in DMEM containing 50% glycerol and stored at -70 ° C;
5. Thaw the human amniotic membrane before use, and then cut it into small pieces with a diameter of about 2 mm after washing with PBSA;
6. Digest the tissue block with trypsin/EDTA at 37 ° C for 30 min;
7. Scrap the digested human amniotic membrane with a plastic scraper to remove the epithelial cells, taking care not to damage the basement membrane;
8. Wash the exposed amniotic membrane with PBSA and adhere its basement membrane surface (the surface of the epithelial cell removed) to the Millicell microporous membrane tissue culture insert;