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Chapter 4 Protocol Building Blocks
4
Wash cells and buffer exchange
(continued)
1
1
Media selection
Depending on the cell type, sensitivity to processing and the properties of the wash media, the rapid
exposure to the new media in counter flow centrifugation can contribute to higher than expected cell
losses. Washing with the same media, culture media (e.g. CTS
HSA, can significantly improve cell recovery.
Note: Forming a fluidized bed in dense media requires higher G force and/or lower flow rates
compared to lighter media. The bed will also take longer to form, especially if the cells are small.
Wash to similar density media
If the wash media has very similar density to the original media, washing can often be conducted at the
loading speed or slightly higher G force (or lower flow rate) with minimal disruption of the fluidized bed.
Wash to more dense media
If the wash media is substantially more dense or higher viscosity than the original media, cells can be
slow to settle. There is also a risk that the denser wash media will not mix with the original media as
it tries to pass through the fluidized bed, instead pushing the bed of cells out of the CFC Chamber.
Sometimes this issue can be addressed by washing at high G and very low flow rates.
72
Image
CTS
™
Rotea
Pump new wash media through the fluidized bed
displacing the original media.
Note: The amount of original media remaining
(residual) is dependent on the volume of new wash
media that passes through the bed but 3 to 5 CFC
Chamber volume changes (e.g. 30 to 50 mL) is often
adequate.
™
™
AIM‑V
™
Counterflow Centrifugation System Process Design User Guide
Details
Medium) or PBS with 1–2%
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