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recombinant FGF2-G3 145 aa protein (Qk052)

FGF2-G3 (145 aa) protein is a thermostable engineered form of FGF-2 (bFGF). Qk052 comprises the 145 aa form of FGF-2 (Qk025) with the nine amino acid substitutions developed by Dvorak et al. 2018. This increases the functional half-life of the protein from <10 h (wild-type) to >7 days (FGF2-G3 145 aa).

FGF2-G3 is used in B8 media (Kuo et al. 2019) for weekend free, high homogeneity induced pluripotent stem cell culture. This new 145 aa thermostable FGF-2 protein allows direct comparison of efficacy in systems using FGF-2 145 aa (Qk025) for chemically defined stem cell and organoid culture media, and cultured meat media development.

High purity 16 kDa bioactive FGF2-G3 145 aa protein. Animal-free (AOF), carrier protein-free and with no His-tag.

Orders are typically shipped same or next day (except Friday).
Easy world-wide ordering, direct or through our distributors.

5010050010005000
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buy online with secure credit card or purchase order
enquiries, bulk and stock reservation, please email orders@qkine.com

1mg may be despatched as 2 x 500µg

summary

  • high purity thermostable FGF2-G3 protein comprising 145 aa form of FGF-2 (Uniprot: P09038) with nine stabilizing amino acid substitutions 

  • 16 kDa

  • expressed in E. coli. 

  • animal-free (AOF) and carrier protein-free.

  • manufactured in our Cambridge, UK laboratories

  • frozen in PBS, 1mg/ml

  • prepare single use aliquots, add carrier protein if desired, store frozen at -20 oC or -80 oC
handling and storage FAQ

featured applications

  • expansion of induced pluripotent, embryonic and mesenchymal stem cells

  • cell expansion

Basic fibroblast growth factor, bFGF, FGF-β, FGF2, Fibroblast growth factor-basic, HBGF-2, FGF2-G3, FGF2-STAB

species neutral

alternative

FGF-2 (145aa) (Qk025): wild-type form of this protein
FGF-G3 (Qk053): the 154 aa form of hyperstable FGF2-G3
FGF-2 154aa (Qk027)
zebrafish FGF-2 (Qk002)

bioactivity

Human LIF Qk036 protein bioactivity lot #14293

FGF2-G3 (145 aa) activity is determined using the Promega serum response element luciferase reporter assay (*) in transfected HEK293T cells. EC50 = 101 pg/ml (6.2 pM).

Cells are treated in triplicate with a serial dilution of FGF2-G3 (145 aa) for 3 hours. Firefly luciferase activity is measured and normalized to the control Renilla luciferase activity. Data from Qk052 lot #104339.

*Promega pGL4.33[luc2P/SRE/Hygro] #E1340

purity

Human LIF Qk036 protein purity SDS-PAGE lot #14293

FGF2-G3 (145 aa) migrates as a major band at 16 kDa in non-reducing (NR) conditions.  A small amount of dimer can be seen (higher molecular weight band). Upon reduction (R), only the 16 kDa band is visible.  No contaminating protein bands are present.

Purified recombinant protein (3 µg) was resolved using 15% w/v SDS-PAGE in reduced (+β-mercaptothanol, R) and non-reduced (NR) conditions and stained with Coomassie Brilliant Blue R250.  Data from Qk052 batch #104339.

further quality assays

  • mass spectrometry: single species with expected mass

  • analytical reversed-phase: single sharp peak 

  • endotoxin: <0.005 EU/μg protein (below level of detection)

  • recovery from stock vial >95%

request CoA/animal-free certificate
download MSDS

We are a company founded and run by scientists to support innovation in stem cell biology and regenerative medicine.  To enhance reliability and reproducibility in your applications, all our products are exceptionally high purity, with complete characterisation and bioactivity analysis on every lot.

Wild-type FGF-2 (145 aa) and FGF-G3 (145 aa) have equivalent bioactivity

FGF2-G3 (145 aa) activity is determined using the Promega serum response element luciferase reporter assay (*) in transfected HEK293T cells.
Cells are treated in triplicate with a serial dilution of FGF2-G3 for 3 hours. Firefly luciferase activity is measured and normalized to the control Renilla luciferase activity.

*Promega pGL4.33[luc2P/SRE/Hygro] #E1340

‘After a long day running samples and analysing data, it was very exciting to see just how stable the FGF2-G3 proteins were. They clearly have more endurance than me!’ – Kerry, Qkine Senior Scientist

Hyperstable FGF2-G3 (145 aa) retains activity pre-incubation with conditioned media at 37oC

+24 hours
+48 hours
+72 hours

FGF2-G3 145 aa (Qk052)            WT FGF2 145 aa (Qk025)

WT FGF2 (Qk025)  and FGF2-G3 (Qk052) were diluted in conditioned media and incubated at 37oC. Samples were taken at daily intervals, serially diluted and then assayed in triplicate for activity using the Promega serum response element luciferase reporter assay (*) in transfected HEK293T cells. Firefly luciferase activity was measured and normalized to the control Renilla luciferase activity (F/R).

FGF2-G3 protein background

the history of FGF2 and stem cell culture

FGF-2 (also known as basic FGF or bFGF) is an essential growth factor for maintaining human embryonic stem cell (hESC) and induced pluripotency stem cell (iPSC) pluripotency in feeder-free and chemically defined stem cell media. It is a core component of widely adopted media including mTESR (Ludwig et al. 2006), StemPRO (Wang et al. 2007) and E8  (Chen et al. 2011). However, FGF-2 is inherently unstable and prone to proteolytic degradation and aggregation. This fundamental biochemical instability, and therefore low half-life in culture media (<10 h), is an important contribution to the need for frequent media changes and challenges in improving homogeneity during stem cell proliferation and subsequent differentiation.

To improve the stability of FGF-2 for stem cell media and regenerative medicine applications, Dvorak and colleagues at Masaryk University used computer-assisted protein engineering to identify an optimal set of nine amino acid substitutions that stabilize FGF-2. These substitutions were designed to avoid structural changes to the FGF receptor 1 (FGFR1) and FGF receptor 2 (FGFR2) binding sites. This thermostable FGF-2 is known as FGF2-G3, or FGF2-STAB® (Dvorak et al. 2017). The biological activity of wild-type FGF-2 is <50% after 10h incubation with conditioned media. In contrast, no reduction in FGF2-G3 biological activity is observed after >7 days incubation with conditioned media at 37oC. Both FGF2-G3 and wild-type FGF-2 maintain hESC pluripotency and expression of pluripotency markers Oct-4 and nanog with equivalent efficacy (Dvorak et al. 2017).

In 2020, Paul Burridge and colleagues at Northwestern University, Chicago, published a protocol for B8 media. This iPSC maintenance media uses thermostable FGF2-G3, along with optimization of media component concentration and composition to reduce media cost and facilitate weekend-free stem cell culture regimes. (Kuo et al. 2020 and updated in Lyra-Leite et al. 2020).

To manufacture and provide FGF2-G3 for stem cell culture, including use in B8 media and emerging applications such as cultured meat, Qkine has licensed the patented FGF2-G3 technology from Enantis/Masaryk University.  We have combined the excellent science behind the FGF2-G3 technology with our protein manufacture expertise to provide gold standard protein for use in cell culture media.  We have removed His tags present in academic forms of the protein, as these may give rise to issues for scientists translating discoveries to the clinical or scale-up.  His tags also introduce unnecessary scientific uncertainty.

Two forms of FGF-2 are used frequently in stem cell culture: 145 aa (Qk025) and 154 aa (Qk027).

To allow scientists to compare directly with their existing FGF-2, we have introduced the nine amino acid substitutions into the 145 aa form of FGF-2 to produce Qk052 FGF2-G3 (145 aa).  The EC50 of the two forms is equivalent and the half-life is extended from less than 1 day to >3 days.

key papers

Dvorak et al. 2018

development of FGF2-G3 form using AI

Koledova et al. 2019

assessment of FGF2-G3 stability and signalling

Kuo et al. 2020

development of B8 media – low cost, weekend free iPSC media

Lyra-Leite et al. 2021

updated protocol for B8 media

Benington et al. 2020

nice review of FGF-2 stability and stabilization methods

Our products are for research use only and not for diagnostic or therapeutic use.  Products are not for resale.

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