Recombinant zebrafish FGF2, bFGF, protein (Qk002)
Fibroblast growth factor 2 (FGF-2) or basic FGF has a broad range of physiological roles including regulation of cell proliferation and differentiation. FGF2 is used to support the maintenance of human embryonic stem cells and proliferation and differentiation of induced pluripotent and mesenchymal stem cells. This version of FGF-2 is used by the core facility at the Cambridge Stem Cell Institute, UK and historically by many of the stem cell groups based at University of Cambridge. FGF-2 or basic FGF from zebrafish has been used extensively to support the maintenance of human stem cells (1–3).
Summary: Qk002 mature domain of zebrafish Danio rerio FGF2, basic FGF (residues 2-154, Uniprot: B3DGE3) expressed in E.coli and purified to homogeneity as described in Ludwig et al. (2).
Form: protein is provided in PBS without carrier protein at 2 mg/ml.
Molecular mass: ~17 kDa
Thaw the sample on ice, spin briefly and dilute with PBS as needed. Our protein are supplied carrier-protein free. If compatible with your work, add carrier protein of your choice such as BSA, HSA or gelatin to further minimise loss by adsorption. Spin in a microfuge for 5 minutes at maximum speed, and divide the solution into suitable aliquots and store at -80°C. We recommend that single-use aliquots should be prepared to avoid freeze-thaw cycles.
Every effort is made to ensure samples are sterile however we recommend sterile filtering after dilution in media or the final working solution.
1. Ludwig, T. E. et al. Derivation of human embryonic stem cells in defined conditions. Nat. Biotechnol. 24, 185–187 (2006).
2. Ludwig, T. E. et al. Feeder-independent culture of human embryonic stem cells. Nat. Methods 3, 637–646 (2006).
3. Chen, G., Gulbranson, D. R., Yu, P., Hou, Z. & Thomson, J. A. Thermal stability of fibroblast growth factor protein is a determinant factor in regulating self-renewal, differentiation, and reprogramming in human pluripotent stem cells. Stem Cells 30, 623–30 (2012).
Result: Result: FGF2 migrates as major band at 17 kDa in non-reducing (-βME) conditions and upon reduction (+βME). The higher molecular mass band at 35 kDa is a dimer that we always see in our highly purified zebrafish FGF2, the presence of this does not affect biological activity.
Purified recombinant protein (7 µg) was resolved using 15% w/v SDS-PAGE in reduced (+β-mercaptothanol, R) and non-reduced conditions (NR) and stained with Coomassie Brilliant Blue R250.
Result: calculated molecular mass of FGF2 is 17036 Da. Molecular mass from this analysis is 17035 Da, consistent with the calculated mass. The calculated molecular mass of dimeric protein is 34064 Da and this is represented in the mass spec with average molecular mass 34062 Da.
MALDI mass spectrometric analysis is used to confirm the molecular mass of the intact protein and to reveal any heterogeneity that would not be evident in SDS-PAGE analysis. The results are compared with calculated mass of the protein with the assumption that all the cysteines are disulphide-linked. The different peaks represent different charge states of the protein.
Result: UV spectrum shows full recovery of protein following aliquoting and lyophilisation.
Absorbance at 280 nm: average 0.093
Recovered concentration: 0.093 cm-1 x 20 / 0.935 cm-1 mg ml-1 = 1.99 mg / ml
The sample was diluted 1:10 in 100 mM sodium phosphate pH 7.4 and the UV spectrum 340-220 nm measured in duplicate. Concentration was calculated using extinction coefficient at 280 nm
Result: Endotoxin level <0.01 EU/ug protein
Stem cell cultures are sensitive to endotoxins1, which can be present in media, serum and as a contaminant on plasticware. We optimise our protein production processes to ensure the lowest possible levels of endotoxin contamination. Our endotoxin pass criteria are set at the industry leading <0.1 EU per ug protein and we aim for <0.01 EU per ug protein. Endotoxin levels in our proteins are determined by an external expert microbiological testing services provider.
1. A biological study establishing the endotoxin limit for in vitro proliferation of human mesenchymal stem cells (2017). Yusuke Nomura, Chie Fukui, Yuki Morishita, Yuji Haishima. Regenerative Therapy, 7, 45-51.
View full batch quality testing data for Qk002
Batch #010, #011
All our proteins are produced in our Cambridge, UK, labs. We provide detailed quality data for each batch because we believe reliable, high quality cytokines and growth factors are critical for successful stem cell and organoid culture.
When we test our proteins, we choose a vial at random and reconstitute as recommended to ensure we are testing as close to the protein you will receive as possible. Biochemical identity and purity is checked using SDS-PAGE, mass spectrometry and analytical reverse phase chromatography. Bioactivity is determined using an appropriate cell-based assay. As stem cells are sensitive to endotoxin levels, we use a high resolution test to ensure endotoxin levels are at industry leading low levels (<0.01 EU per µg protein). We also check that the correct amount of protein is recovered from the vial – it might sound basic but if you order 100 µg, we believe you should receive 100 µg so when you use the proteins you can rely on your calculated dilution.
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Order online and upload your PO or pay by credit card, whichever is easiest for you, or email your PO to email@example.com.
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Recombinant mature domain of human Activin A protein (residues 311-426, Uniprot: P08476) expressed in E.coli (animal-derived component free). Protein supplied lyophilized with no carrier protein present. Activin A is used in stem cell maintenance and differentiation.