Can you provide a protocol for the expansion of HPCs using PromoCell HPC Expansion Medium XF?
Yes. Below is a short, standardized protocol for expanding HPCs using HPC Expansion Medium XF supplemented with Cytokine Mix E.
Short protocol
1. Thawing cryopreserved HPCs
Thaw the cells for 2 minutes at 37°C in a water bath.
Transfer the cells into 9 ml complete HPC Expansion Medium XF (supplemented with Cytokine Mix E) and determine the cell count.
2. Initial seeding
Centrifuge for 10 minutes at 240×g.
Aspirate the supernatant and resuspend the pellet at 10,000 cells/ml in complete HPC Expansion Medium XF.
Seed the cells into an appropriate suspension culture vessel (e.g. 20,000 cells in 2 ml cell suspension per well of a 6‑well plate) and incubate for 2–3 days at 37 °C and 5% CO₂.
Alternatively, plate freshly isolated HPCs at a density of 10,000 cells/ml in cytokine-supplemented complete medium in a suitable suspension culture vessel.
3. Cell expansion
After 2-3 days, add fresh complete medium to double the culture volume (e.g. add 4 ml fresh medium to 4 ml culture, resulting in a total volume of 8 ml).
Continue incubation for 10-12 days.
Perform a partial medium change every 2-3 days
Example: If the culture volume is 8 ml, discard 4 ml after centrifugation; resuspend the cells in the remaining 4 ml, then add to 12 ml fresh complete medium (final 16 ml).
5. Harvest expanded HPCs (day 13-15)
For more details, please refer to the corresponding Application Note.
Short protocol
1. Thawing cryopreserved HPCs
Thaw the cells for 2 minutes at 37°C in a water bath.
Transfer the cells into 9 ml complete HPC Expansion Medium XF (supplemented with Cytokine Mix E) and determine the cell count.
2. Initial seeding
Centrifuge for 10 minutes at 240×g.
Aspirate the supernatant and resuspend the pellet at 10,000 cells/ml in complete HPC Expansion Medium XF.
Seed the cells into an appropriate suspension culture vessel (e.g. 20,000 cells in 2 ml cell suspension per well of a 6‑well plate) and incubate for 2–3 days at 37 °C and 5% CO₂.
Alternatively, plate freshly isolated HPCs at a density of 10,000 cells/ml in cytokine-supplemented complete medium in a suitable suspension culture vessel.
3. Cell expansion
After 2-3 days, add fresh complete medium to double the culture volume (e.g. add 4 ml fresh medium to 4 ml culture, resulting in a total volume of 8 ml).
Continue incubation for 10-12 days.
Perform a partial medium change every 2-3 days
Example: If the culture volume is 8 ml, discard 4 ml after centrifugation; resuspend the cells in the remaining 4 ml, then add to 12 ml fresh complete medium (final 16 ml).
5. Harvest expanded HPCs (day 13-15)
For more details, please refer to the corresponding Application Note.
