How to transfer iPS cells to DEF-CS
How to transfer iPS cells to DEF-CS
Cells grown in other culture systems:
Undifferentiated human iPS cells maintained in other culture systems can be readily transferred to the DEF-CS system. It takes between 2 and 5 passages to adapt a cell line to the DEF-CS system. The standard DEF-CS culture protocol should be followed, although some modifications may increase the success of transfer:
The cells may benefit from a higher concentration of DEF-CS COAT-1 (coating substance). Use a dilution of 1:10 or 1:5 during the first passage to provide extra support during the transfer process.
A higher seeding density for the first passage (e.g., 8.0 x 104 cells/cm2) can also be beneficial to the cells.
Newly transferred cells might initially grow at a slightly slower rate. A suitable passage interval might therefore be between 3 and 7 days for the first passages. The cells are ready for passage when they have acquired the morphology in the images above (see 1.5 x 105 cells/cm2 and 2.0 x 105 cells/cm2). Even if the cells are sparse after 7 days in culture, passaging is still recommended.
Fresh iPS cell cultures:
Fresh cultures can be transferred to the DEF-CS system at passage. The cells should be dissociated according to the protocol of the previous system, and seeded as single cells or aggregates according to the DEF-CS protocol using a 1:1 split ratio.
Frozen iPS cells:
Cryopreserved cells can be thawed directly into the DEF-CS system.