PFHM-II Protein-Free Hybridoma Medium (liquid) - FAQs

How do I adapt my cells to serum-free medium?

Cells can be adapted by Sequential or Direct Adaptation. Suggested protocols for each are below, and you can also find more information by searching "Adaptation of Cell Cultures to a Serum-Free Medium" from our website home page.

SEQUENTIAL ADAPTATION
1) Subculture the cells growing in serum-supplemented medium into a 25%:75% mixture of SFM and serum supplemented medium.
2) When the cell density is 5 x 10E5 cells/ml, subculture the cells into a 50%:50% mixture of SFM and serum supplemented medium at a cell density 2.5 x 10E5 to 3 x 10E5 cells/ml.
3) Continue to subculture after the cell density 5 x 10E5 cells/ml in gradually increasing proportions of SFM until the serum is ~0.1% with about 85% cell viability.
4) Subculture the cells into SFM with an innoculum of 2.5 x 10E5 to 3 x 10E5 cells/ml.
5) When the cell density is 1 x 10E6 to 3 x 10E6 cells/ml (4 to 6 days post planting) subculture the cells again.
6) Stock cultures of SFM adapted cells should be subcultured in SFM every 3 to 5 days when the cell density is 1 x 10E6 to 3 x 10E6 cells/ml with 90% viability.

DIRECT ADAPTATION
Some cells can be directly adapted from serum-containing medium to SFM. For direct adaptation, the cell innoculum should be 1.5 x 10E5 to 3 x 10E5 cells/ml.
Cells should be subcultured when the cell density is 1 x 10E6 to 3 x 10E6 cells/ml. Cells are fully adapted to SFM when the cell density is 2 x 10E6 to 4 x 10E6 cells/ml after 4 to 7 days in culture.
Stock cultures of cells adapted to SFM should be subcultured in SFM every 3 to 5 days when the cell density is 1 x 10E6 to 3 x 10E6 cells/ml with 90% viability.

Why is it necessary to gradually adapt the cells to serum-free medium?

Some cells, such as insect cells, are sensitive to changes in their medium. By sequentially adapting cells, the medium is changed with minimal effects on cell growth.

Find additional tips, troubleshooting help, and resources within our Cell Culture Support Center.

Can I freeze PFHM-II Protein-Free Hybridoma Medium (liquid) (Cat. No. 12040077)?

We do not recommend freezing our liquid media products. Freezing can cause some components like amino acids and inorganic salts to precipitate upon thawing.

Find additional tips, troubleshooting help, and resources within our Cell Culture Support Center.