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Related Concept Videos

Induced Pluripotent Stem Cells01:13

Induced Pluripotent Stem Cells

Stem cells are undifferentiated cells that divide and produce different types of cells. Ordinarily, cells that have differentiated into a specific cell type are post-mitotic—that is, they no longer divide. However, scientists have found a way to reprogram these mature cells so that they “de-differentiate” and return to an unspecialized, proliferative state. These cells are also pluripotent like embryonic stem cells—able to produce all cell types—and are therefore called induced pluripotent stem...

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Methods for Automated Single Cell Isolation and Sub-Cloning of Human Pluripotent Stem Cells.

Valeria Fernandez Vallone1,2, Narasimha Swamy Telugu3,4, Iris Fischer1,2

  • 1Charité-Universitätsmedizin Berlin, Berlin, Germany.

Current Protocols in Stem Cell Biology
|September 21, 2020
PubMed
Summary

Automated cell isolation devices improve human pluripotent stem cell (hPSC) cloning efficiency and precision. These automated workflows offer a robust alternative to manual methods for high-throughput hPSC clonal selection and expansion.

Keywords:
automationhPSCkaryotypingsingle cell isolationsub-cloning

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Area of Science:

  • Stem Cell Biology
  • Biotechnology
  • Genomics

Background:

  • Human pluripotent stem cells (hPSCs) are crucial for disease modeling, drug discovery, and regenerative medicine.
  • Traditional hPSC single-cell cloning methods (limiting dilution, manual picking) are inefficient, labor-intensive, and lack proof of clonality.
  • CRISPR/Cas9 genome editing and cell-based therapies require precise clonal expansion from single hPSCs.

Purpose of the Study:

  • To evaluate automated cell isolation and dispensing devices for enhancing hPSC single-cell cloning.
  • To establish optimized protocols for automated hPSC clonal expansion using different platforms.
  • To demonstrate the maintenance of pluripotency and genetic stability in automated hPSC clones.

Main Methods:

  • Application of three automated cell isolation and dispensing devices: iotaSciences IsoCell, Cellenion CellenONE, and Cytena single-cell dispenser.
  • Optimization of cell culture conditions and specific protocols for each device.
  • Assessment of single cell-derived hPSC clones for pluripotency, genetic stability, and karyotypic integrity.

Main Results:

  • Automated devices significantly improve the efficiency and precision of hPSC single-cell cloning compared to manual methods.
  • Established workflows ensure the maintenance of pluripotency and genetic stability in expanded hPSC clones.
  • The automated approach successfully identified karyotypic mosaicism in bulk hPSC cultures.

Conclusions:

  • Automated cell isolation platforms provide a robust and efficient solution for high-throughput hPSC clonal selection and expansion.
  • These workflows are essential for advancing hPSC applications in research and therapeutics.
  • The described methods facilitate the generation of genetically stable and pluripotent hPSC clones.