Late-adhering human embryonic stem cell clumps during serial passage can yield morphologically 'normal-looking'...

Area of Science:

• Stem Cell Biology
• Developmental Biology
• Cell Culture Techniques

Background:

• Serial passage of human embryonic stem (hES) cells involves discarding non-adherent cell clumps.
• Routine discarding may lead to the loss of viable hES cells with potential for differentiation.

Purpose of the Study:

• To investigate the viability and differentiation potential of non-adhered hES cell clumps during serial passage.
• To determine if retaining late-adhering hES cell clumps impacts overall culture quality.

Main Methods:

• Observation of hES cell clump morphology after overnight culture on murine embryonic fibroblast (MEF) feeder layers.
• Assessment of adherence and differentiation potential of non-adhered clumps after further culture.
• Comparison of colony morphology between retained and discarded hES cell clumps.

Main Results:

• Viable, non-adhered hES cell clumps with rounded, sharp edges adhered within 24 hours and formed morphologically normal, undifferentiated colonies.
• Non-adherent clumps with jagged edges were non-viable or represented senescent MEF and did not adhere.
• Late-adhering hES cell clumps, even those initially clumped with other colonies, yielded undifferentiated colonies comparable to their neighbors.

Conclusions:

• Retaining late-adhering hES cell clumps during serial passage is a potentially valuable strategy.
• This method could prevent the loss of viable stem cells and maintain culture efficiency.
• Morphological assessment of non-adhered clumps can distinguish viable stem cells from non-viable debris.