Stem cell maintenance

TGF beta 1 PLUS comparison

Maintenance of pluripotency in human iPSC and ESC

Several standard defined media have been adopted for human stem cell maintenance including mTESR, E8 and StemPro. Most recently, weekend-free media B8 developed by Paul Burridge at Northwestern University, utilises thermostable (heat-stable) FGF-2 (FGF2-G3) developed by Dvorak and colleagues at Masaryk University and Enantis Ltd.

In the search for conditions that maintain pluripotency in human pluripotency stem cells, several media formulations have been described (the composition of most popular of which are shown in table 1), in general each of which targets 3 main signalling pathways.1

  • Activation of the PI3K/AKT/mTOR and MAPK/ERK pathways by FGF-2 and/or neuregulin 1 (NRG-1), which bind FGFR1/FGFR4 or ERBB3/ERBB4
  • Activation of the TGF-β signaling pathway by TGF-β1, nodal, or activin A, which bind TGFBR1/2 and/or ACVR2A/2B/1B/1C (nodal is used less commonly in pluripotent medium formulations due to the expression of the nodal antagonists LEFTY1/2 in hPSCs)
  • Activation of the PI3K/AKT pathway, promoting cell survival and growth, by insulin or insulin growth factor (IGF-1), which binds INSR and IGF1R

Products for iPSC and ESC maintenance media

mTESR and E8

TGF-B1 PLUS (Qk010)
FGF-2 / bFGF 145 aa (Qk025)
FGF-2 / bFGF 154 aa (Qk027)
zebrafish FGF-2 /bFGF (Qk002)

StemPro media

TGF-B1 PLUS (Qk010)
TGF-B3 (Qk054)
heregulin beta 1 / NRG-1 (Qk045)
FGF-2 / bFGF 145 aa (Qk025)
FGF-2 / bFGF 154 aa (Qk027)
zebrafish FGF-2 /bFGF (Qk002)
activin A (Qk001)
IGF-1 (Qk047)
IGF-1 LR3 (Qk041)

B8 media

heat stable FGF2-G3 / bFGF / FGF2-STAB® 154 aa (Qk053)
heat stable FGF2-G3 / bFGF / FGF2-STAB® 145 aa (Qk052)
TGF-B1 PLUS (Qk010)
TGF-B3 (Qk054)
heregulin beta 1 / NRG-1 (Qk045)
FGF-2 / bFGF 145 aa (Qk025)
FGF-2 / bFGF 154 aa (Qk027)
zebrafish FGF-2 /bFGF (Qk002)
IGF-1 (Qk047)
IGF-1 LR3 (Qk041)

Table 1. the composition of four of the most popular human pluripotency media

ComponentsRecipe
mTeSR1DMEM/F12, BSA, FGF2, TGFβ1, insulin, transferrin, cholesterol, lipids, pipecolic acid, GABA, b-mercaptoethanolLudwig et al 20062
E8DMEM/F12, FGF2, TGFβ1, insulin, transferrin, selenium, ascorbic acidBeers et al 20123
B8DMEM/F12, FGF2-G3 (FGF2-STAB®), TGFβ3/TGFβ1, NRG1 (Heregulin-β1), insulin/IGF1 LR3, ascorbic acid, transferrin, sodium selenite, sodium bicarbonateLyra-Leite et al 20214
StemProDMEM/F12, BSA, FGF2, TGFβ1, Activin A, IGF-1 LR3, NRG1 (Heregulin-β1), transferrin,Wang et al 20075

Product focus: enhanced pluripotency maintenance using animal-free TGF beta 1 PLUS

Human/bovine/porcine TGF-β1 PLUS protein is the first entirely animal-free recombinant human transforming growth factor beta 1 (TGF-β1) protein for highly reproducible results and compatible with chemically-defined stem cell media. TGF-β1 is used extensively in E8-style induced pluripotent (iPSC) and embryonic (ESC) stem cell media.

High purity 24 kDa dimer comprising optimised mature domain of TGF-β1 protein, animal-free (AF) and carrier-protein free (CF). Our TGF-β1 PLUS protein has been extensively tested for maintenance of iPSC pluripotency by the specialist stem cell biotechnology company, Stemnovate, Cambridge, UK

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References

  1. Kuo, Hui-Hsuan, et al. ‘Negligible-Cost and Weekend-Free Chemically Defined Human IPSC Culture’. Stem Cell Reports, vol. 14, no. 2, Feb. 2020, pp. 256–70.
  2. Ludwig, Tenneille E., et al. ‘Feeder-Independent Culture of Human Embryonic Stem Cells’. Nature Methods, vol. 3, no. 8, Aug. 2006, pp. 637–46.
  3. Beers, Jeanette, et al. ‘Passaging and Colony Expansion of Human Pluripotent Stem Cells by Enzyme-Free Dissociation in Chemically Defined Culture Conditions’. Nature Protocols, vol. 7, no. 11, 2012, pp. 2029–40.
  4. Lyra-Leite, Davi Marco, et al. ‘An Updated Protocol for the Cost-Effective and Weekend-Free Culture of Human Induced Pluripotent Stem Cells’. STAR Protocols, vol. 2, no. 1, Mar. 2021, p. 100213.
  5. Wang, Linlin, et al. ‘Self-Renewal of Human Embryonic Stem Cells Requires Insulin-like Growth Factor-1 Receptor and ERBB2 Receptor Signaling’. Blood, vol. 110, no. 12, Dec. 2007, pp. 4111–19.
  6. Chen, Chen-Yun, et al. ‘Mechanisms of Pluripotency Maintenance in Mouse Embryonic Stem Cells’. Cellular and Molecular Life Sciences, vol. 74, no. 10, May 2017, pp. 1805–17.
  7. Merrill, B. J. ‘Wnt Pathway Regulation of Embryonic Stem Cell Self-Renewal’. Cold Spring Harbor Perspectives in Biology, vol. 4, no. 9, Sept. 2012, pp. a007971–a007971.