Rat Mesenchymal Stem Cells (MSCs) Isolation and Culture Procedures
1. Kill 3-4 6-weeks old Wistar rats with CO2.
2. Harvest the
femurs aseptically.
3. Completely remove the adherent soft tissues.
4. Exercise he proximal and distal ends of the femur at a level
just into the beginning of the marrow cavity.
5. Flush the bone marrow cavity with a 18-gauge needle with alpha-MEM medium enriched
with antibiotics
and antimycotic solution, to get the bone marrow plugs.
6. Pass through 18 and 22-gauge
needles to disperse the bone marrow plugs to get single cell suspension.
7. Centrifuge and resuspend the cells with 20% FBS
alpha-MEM medium enriched with antibiotics
and antimycotic solution.
8. Disrupt the red blood cells with acetic
acid soultion.
9. Wash with PBS.
10.Plate nucleated cells in a 75 ml flask with 20% FBA alpha-MEM medium, and incubate at 37
centigrade in a humidified incubator of 95% and 5% CO2.
11.Change half of the full medium after 48 hours.
12. Digest the cells with
trypsin-EDTA solution and passage the cells, when they are 80-90% confluence.
13. Evaluate MSCs markers.
Protocols
- Culture
of Human Embryonic Stem Cells (hESC)
- Establishment and characterization of human embryonic stem cell line
- Rat stem cell isolation
and culture
- Mouse stem cell isolation and culture
- Induction of Stem Cell Differentiation
- Cancer stem
cells
- Video Data on Stem Cell Research (click to show)