NaïveCult™ Induction Kit

Defined medium kit for transgene-free induction of reset naïve human ES and iPS cells
概要
NaïveCult™ Induction Kit contains defined cell culture media for the chemical induction of transgene-free reset naïve human embryonic stem (ES) and induced pluripotent stem (iPS) cells from the primed state. The kit was developed under license from the Cambridge Stem Cell Institute at the University of Cambridge, UK based on Guo et al. (2017). This kit works robustly across multiple primed human ES and iPS cells maintained in mTeSR™1 (Catalog #85850) or TeSR™-E8™ (Catalog #05990). Reset naïve human pluripotent stem cells (hPSCs) generated using NaïveCult™ Induction Kit demonstrate uniformly domed, phase-bright colonies with refractive edges and maintain gene expression profiles associated with reset naïve hPSCs (Guo et al. 2016; Guo et al. 2017; Takashima et al.).

NaïveCult™ Induction Kit requires a histone deacetylase inhibitor (HDACi) that is sold separately (either Sodium Butyrate [Catalog #72242] or Valproic Acid [Catalog #72292]).

References
Takashima Y et al. (2014) Cell 158(6): 1254–69.
Guo G et al. (2016) Stem Cell Reports 6(4): 437–46.
Guo G et al. (2017) Development 144(15): 2748–63." 920
Advantages
• Transgene-free chemical induction of reset human naïve ES and iPS cells
• Robust induction across multiple human ES and iPS cell lines
• Generates hPSCs with high expression of naïve-associated genes
• Defined formulation containing high-quality, pre-screened components
Components
  • NaïveCult™ Induction Basal Medium, 76 mL
  • NaïveCult™ 20X Induction Supplement A, 4 mL
  • NaïveCult™ 1000X Induction Supplement B, 80 µL
  • NaïveCult™ Expansion Basal Medium, 2 x 400 mL
  • NaïveCult™ 5X Induction Supplement C, 200 mL
  • NaïveCult™ 1000X Induction Supplement D, 500 µL
  • NaïveCult™ 1000X Expansion Supplement, 500 µL
Subtype
Specialized Media
Cell Type
Pluripotent Stem Cells
Species
Human
Application
Cell Culture, Expansion, Maintenance
Brand
NaïveCult
Area of Interest
Stem Cell Biology
数据及文献

Data

Figure 1. Schematic of Reversion of Primed to Naïve hPSCs

Primed hPSCs, grown either in mTeSR™1 or TeSR™-E8™ on Corning® Matrigel®, are plated as single cells on inactivated murine embryonic fibroblasts (iMEFs) and treated with Rho-kinase inhibition (10 μM Y-27632) for 24 hours in hypoxic conditions. On day 1, medium is changed to Induction Medium 1 supplemented with a histone deacetylase inhibitor (HDACi) and cells are cultured for 3 days with daily medium changes. On day 4, medium is changed to Induction Medium 2 and cells are cultured until day 11 with daily medium changes. On day 11, hPSCs are passaged as single cells onto fresh iMEFs and subsequently passaged every 3 - 4 days until the end of passage 2 in Induction Medium 2. From passage 3, hPSCs are cultured in Induction Medium 3. Background differentiation will decrease between passage 3 and 8. At this time cells can be transferred into NaïveCult™ Expansion Medium for long-term maintenance and expansion.

Figure 2. Human ES and iPS Cells Can Be Reverted to a Naïve State

Representative images of human (A) passage-7 H9 ES cells and (B) passage-9 WLS-1C iPS cells that were reverted to a naïve state using the NaïveCult™ Induction Kit and subsequently cultured in NaïveCult™ Expansion Medium. During reversion, colonies change from a flat morphology to a tightly packed, domed morphology with refractive edges characteristic of naïve-state hPSCs.

Figure 3. hPSCs Cultured in the NaïveCult™ Media System Express High Levels of Factors Associated with Naïve hPSCs

Gene expression profile of human (A) H9 ES cells and (B) WLS-1C iPS cells reverted using the NaïveCult™ Induction Kit and maintained in NaïveCult™ Expansion Medium.

Figure 4. Reset Naïve Human PSCs Cultured in NaïveCult™ Expansion Medium are Capable of Tri-Lineage Differentiation Following Re-Priming

Reset naïve human iPS cells cultured in NaïveCult™ Expansion Medium were re-primed in mTeSR™1 and differentiated to all three somatic lineages. (A) Representative flow cytometry plot of human WLS-1C iPS cells differentiated using the STEMdiff™ Definitive Endoderm Kit (Catalog #05110) demonstrating >90% CXCR4+/SOX17+ definitive endodermal progenitors. (B) Representative flow cytometry plot of human WLS-1C iPS cells differentiated using STEMdiff™ Mesodermal Induction Medium (Catalog #05220) demonstrating >80% Brachyury+/OCT4- mesodermal progenitors. (C) Representative immunofluorescence image of PAX6 (green) and SOX1 (red) double positive neural progenitors derived from human WLS-1C iPS cells differentiated using the STEMdiff™ SMADi Neural Induction Kit (Catalog #08581).

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