StemCore passages pluripotent stem cells using both manual (‘cut & paste’ or 'pieces') and enzymatic methods. Each method, described in more detail below, is suitable for particular applications.
Manual passaging (also referred as 'pieces culture')
- Stem cells grow as large, round, clearly defined colonies with a distinctive edge and often develop a differentiated middle 'button'
- Passaging involves manual cutting of individual colonies into fragments and transfer of these fragments to new culture dishes, usually every seven days
- The most commonly used culture vessel used with this method is an Organ Culture Dish (also called IVF-dish)
- Has been linked with the ability to maintain chromosomal stability for prolonged periods
- Exclusion of differentiated cells in each passage allows continuous maintenance of mainly undifferentiated colonies
- Mainly used for maintenance of stock cultures
- Technically demanding and time consuming
Enzymatic passaging as small clumps*
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Stem cells grow as clearly defined colonies of varying size
- Passaging involves digestion of colonies with Collagenase IV, Collagenase I or Dispase to form small clumps, usually every 6-7 days
- Used for stem cell applications where clumps of cells are desired
Enzymatic passaging as single cells*
- Cultures appear initially as small, tightly packed colonies, later almost as a confluent layer of cells
- Passaging involves treatment of cultures with TrypLE™ Select, TrypLE™ Express or Trypsin to form a single cell suspension, usually every 3-4 days
- Used for applications where knowledge of exact cell numbers is important
*Note: Passaging hESCs as single cells or small clumps is thought to increase the risk of chromosomal abnormalities. Hence, enzymatically passaged cultures are not maintained for more than 20 passages.
TrypLE™ Select and TrypLE™ Express are trademarks of Life Technologies.