How do I transfer my cells from feeder cells to the Biolaminin 521 culture substrate?
We recommend transferring the cells as single cells (or as small clumps) and always with the addition of ROCKi for the first few (3-5) passages. Once the cells are adapted to the Biolaminin 521 matrix, the cells can be cultured as single cells without ROCKi. This may take up to 5 passages. If the cells are hard to adapt, use a higher coating concentration (10 µg/ml for LN521 or 20 µg/ml for MX521 and CT521) and seed at a higher cell density 50 000–100 000 cells/cm2. Once the cells are adapted a lower coating and seeding concentration often can be used.
It is important that the cells transferred to the Biolaminin 521 matrix are of high quality. Biolaminin 521 will generally also support differentiated cells so carefully select only undifferentiated cell areas for transfer.
When changing from feeders, the cells might display a different morphology for the first few passages, likely due to the packed monolayer the cells form when confluent, rather than thick colonies as seen on feeders. It is important that the cells are of high quality when being transferred from feeders to the Biolaminin 521 substrate. Biolaminin 521 supports the maintenance of pluripotent cells but may also support some differentiated cells. Therefore, it is important that only undifferentiated cell colonies are being transferred.