Short protocol:
- Thaw the cells (C-12921) for 2 min in a 37°C waterbath. Dilute in 9 ml of complete HPC Expansion Medium XF (+ Cytokine Mix E) and count the cells
- Spin down for 10 min at 240xg, aspirate the supernatant, resuspend the pellet at 20,000 cells/ml HPC Expansion Medium XF
- Plate in an appropriate suspension culture vessel and incubate the culture for 2-3 days at 37°C and 5% CO₂
- Then double the media volume by adding fresh complete medium, e.g., 4 ml suspension culture + 4 ml fresh medium (= 8 ml)
- Incubate the cells for an additional 10-12 days by performing a partial medium change every 2-3 days
Example partial medium change: For a culture volume of 8 ml, spin down the cells, aspirate and discard 4 ml of the supernatant, resuspend the cells and add 12 ml of fresh complete medium (= 16 ml).
In combination with the Cytokine Mix E, the HPC Expansion Medium XF typically promotes a 300-1,000 fold expansion of the total cell population. After 2 weeks of expansion about 20-30% of the population express CD34+, indicating a 50-200 fold expansion of CD34+ progenitor cells.