Scalable passaging of adherent human pluripotent stem cells
Ying Nie1, Patrick Walsh1, Diana L Clarke1
1Cell Therapy Development Services, Lonza-Walkersville, Inc., Walkersville, Maryland, United States of America.
Plos One
|February 6, 2014
Summary
A new non-enzymatic method using sodium citrate efficiently detaches human pluripotent stem cells (hPSCs) for large-scale culture. This method improves cell viability and reduces processing time for clinical applications.
Area of Science:
- Stem Cell Biology
- Biotechnology
- Cell Culture
Background:
- Current methods for passaging human pluripotent stem cells (hPSCs) are labor-intensive and reduce cell viability.
- Existing techniques are not suitable for the large-scale production required for clinical applications.
Purpose of the Study:
- To develop a novel, non-enzymatic method for passaging hPSCs.
- To improve cell viability and enable scalable expansion of hPSCs.
Main Methods:
- Developed a non-enzymatic passaging solution using hypertonic sodium citrate.
- Detached adherent hPSCs as multicellular aggregates with minimal manual intervention.
- Recovered aggregates in calcium-containing medium.
Main Results:
- Achieved efficient and gentle detachment of hPSCs.
- Maintained high post-detachment cell viability (97%±1%).
- Demonstrated successful subculturing for 25 passages with retained pluripotency, normal karyotype, and differentiation potential.
Conclusions:
- The sodium citrate method offers a scalable, efficient, and gentle alternative for hPSC passaging.
- This technique supports the expansion of high-quality hPSC cultures for clinical applications.
- The method preserves critical hPSC characteristics, including pluripotency and genetic stability.
