4-5 days after cell split, when iPSCs reach a confluency of 30-60%, aspirate out spend medium and add 1ml 0.5mM EDTA/PBS. Incubate cells at 37C for ~5min. When colonies start to detach, gently harvest all the cells into a 5-ml or 15-ml or 50-ml tube by pippeting 1-2 times. Avoid multiple pippetting. Breaking down clumps into single cells may substantially decrease cell survival. May add iPSC medium to harvest all the cells. Spin down at 200g for 2-3 minutes. Carefully aspirate out supernatent and gently resuspend cells in 0.5ml iPSC medium. Add 0.5ml freezing medium (TM023) and ROCK inhibitor. Mix well by flipping the vial and transfer the medium to a cryovial. Finally transfer the vial to -80C freezer for short-term storage (days to weeks). After cells are frozen, you may then transfer the vial to liquid nitrogen tank for long-term storage (years).

iPSCs require seeder cells before thawing and plating the iPSCs. Seed ~3x10⁵ feeder cells in each well of a 6-well plate. After thawing the iPSCs, put all the cells from the vial into 1 well. After several days, you may start splitting the iPSCs into multiple wells depending on how many cells survived. The density mentioned above is for the seeder cells which should be at 3x10⁵. Afterwards, please plate the entire vial of iPSCs into one well at 10⁶ concentration.

We only provide clones that have undergone FACS verification for expression of pluripotency markers such as OCT4, NANOG, TRA-1-60. It is possible to assay for additional markers upon request.
We will also only pick from colonies displaying a typical iPSC morphology.