Recombinant human TGF-β1 PLUS protein (Qk010)

£116.00£6,960.00

Recombinant human 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. Our  TGF-β1 PLUS protein has been extensively tested for maintenance of iPSC pluripotency by the specialist stem cell biotechnology company, Stemnovate, Cambridge, UK.

*Benchmark our TGF-β1 PLUS against your existing supplier*

SKU: Qk010_TGFB1 Categories: , Tags: , ,

Recombinant human transforming growth factor-beta 1 (TGF-β1) protein is an essential growth factor in many iPSC and ESC maintenance media, including the commonly used chemically-defined E8-based stem cell culture medias. To date, recombinant human TGF-β1 has only been produced from mammalian cell protein expression systems (HEK or CHO), where endogenous protein contaminants, cost and animal-free status is a challenge.  Our protein experts have optimized TGF-β1 allowing us to produce the first highly bioactive protein in an E. coli fully animal-free expression system. 

As part of our ongoing mission to redefine industry standards for growth factor and cytokine biochemical quality, we’re delighted to introduce the first optimized, truly animal-free recombinant human TGF-β1 for our stem cell maintenance range.

Benefits:

  • Truly animal-free alternative for chemically-defined stem cell culture media
  • High potency in iPSC culture with high pluripotency (Nanog) marker expression
  • High purity, extensive biochemical data and exceptional batch-to-batch consistency

Summary: Qk010 is a modified version of human TGF-β1 protein (Uniprot: P01137), which has been optimized for high purity expression in E. coli. The mature protein is a disulfide-linked dimer.

Molecular mass: ~24 kDa (for the dimer)

Form: Protein is provided lyophilized from a fully volatile solution. Animal-derived component and carrier protein-free.

Transforming growth factor-beta 1, TGF-beta 1, TGFβ1, TGFB1, TGF B1

Example data

Protein purity: SDS-PAGE in reduced and non-reduced conditions

Bioactivity: TGF-β1-responsive luciferase reporter assay

Recombinant human TGF β1 PLUS protein bioactivity

Purity and bioactivity

For peace of mind that our recombinant human TGF-β1 PLUS protein will work exactly the same way every day, from batch to batch and at any scale you need, we conduct extensive purity and bioactivity analysis, including SDS-PAGE, mass spectrometry, UV spectroscopy and endotoxin level testing.

To confirm the biochemical identity of our recombinant human TGF-β1 PLUS protein and ensure that its purity meets our rigorous standards, we conduct SDS-PAGE on every protein batch. As recombinant human TGF-β1 PLUS protein is a disulfide-linked dimer, we use both reduced and non-reduced conditions, as well as appropriate stains for our SDS-PAGE analysis.

We check the final refolded protein is bioactive using a TGF-β1-responsive firefly luciferase reporter assay in HEK293T cells. By knowing what the expected activity of the protein is and measuring calibrant alongside each batch of protein, we can use this bioassay to define a complete dose-response curve and check the EC50 value of the preparation.

In addition, our TGF-β1 PLUS has also been validated for its ability to effectively maintain iPSC pluripotency by the specialist stem cell biotechnology company, Stemnovate, Cambridge, UK.

Find out more about our extensive purity testing in the next tab.

Research applications

iPSC/ESC maintenance

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.  Please contact us with questions any time by email at support@qkine.com or phone +44 (0) 1223 491486 / US toll free 1-866 877 2185.

Order online and upload your PO or pay by credit card,  or email your PO to orders@qkine.com.

We provide bulk orders and stock reservation for sensitive applications, please email us.

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

For final purity and activity tests on our proteins, we choose a vial at random and reconstitute as recommended. Biochemical identity and purity are determined using SDS-PAGE, mass spectrometry and analytical reversed-phase chromatography. Bioactivity is quantified 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.

Purity: SDS-PAGE

Result: TGF-β1 PLUS (Qk010) dimer migrates as a single band at 24 kDa in non-reducing (NR) and 13 kDa as a single monomeric species upon reduction (R) with β-mercaptoethanol. High purity yield of dimeric protein (bioactive form).

Qk010 TGFB1 purity in SDS-PAGE

Purified recombinant protein (7 μg) was resolved using 15% w/v SDS-PAGE in reduced (+β-mercaptoethanol, R) and non-reduced conditions (NR) and stained with Coomassie Brilliant Blue R250. Data from Qk010 batch #012.

Bioactivity: luciferase reporter assay

Result: TGF-β1 PLUS (Qk010) is highly bioactive compared to mammalian-expressed TGF-β1 preparations from other suppliers.

Recombinant TGFB1 PLUS bioactivity in luciferase reporter assay

Comparative activity determined using a quantitative luciferase reporter assay in transiently transfected HEK293T cells. Cells were treated (in triplicate) with a serial dilution of TGF-β1. Firefly luciferase activity is measured and normalized to the control Renilla luciferase activity. TGF-β1 PLUS EC50 1.4 pM. Suppliers 1 and 2 have EC50 ~3.5 pM and 38 pM respectively.

Purity: mass spec analysis

Result: Consistent with the calculated mass. No heterogeneity is present.

Mass spectrometry analysis for recombinant human TGF-β1 PLUS protein

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 disulfide-linked. The different peaks represent different charge states of the protein. Data from Qk010 batch #011.

Purity: endotoxin level determination

Result: Endotoxin level <0.005 EU/µg protein (below level of detection)

Stem cell cultures are sensitive to endotoxins1, which can be present in media, serum and as a contaminant on plasticware. We optimize 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. Example data from Qk010 batch #011.

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.

TGF-β1 PLUS maintains iPSC pluripotency with high Nanog expression

TGF-β1 PLUS is highly effective at maintaining iPSC pluripotency in chemically-defined, serum and feeder-free culture

Nanog Tra1-60 expression in iPSC maintained in TGFB1 PLUS

Immuno-staining for pluripotency markers Tra 1-60 and Nanog show high levels of expression in iPSC lines maintained in TGF-β1 PLUS (Qk010)-containing defined media. In this study, Qkine TGF-β1 PLUS performed better than the TGF-β1 used routinely by Stemnovate.

Phase contrast iPSC maintained in Qk010 TGF beta 1 and Qk025 FGF2 day 3

TGF-β1 PLUS (Qk010) maintains pluripotency and good colony morphology at 1 ng/ml in a chemically-defined serum and feeder-free iPSC culture. TGF-β1 PLUS used at 1 ng/ml; Qk025 FGF2 used at 100 ng/ml.

TGF-β1 PLUS in combination with Qkine FGF2 shows enhanced bioactivity when benchmarked against other suppliers

Comparison of Nanog immunofluorescence in cells maintained in TGF beta 1 from Qkine and supplier 1

Stemnovate IPS media (chemically-defined, serum and feeder-free culture). Higher Nanog pluripotency marker immuno-staining in human iPSC cultured in Qkine FGF2 (Qk025) and TGF-β1 PLUS(Qk010).TGF-β1 PLUS used at 1 ng/ml.

All experiments have been conducted by the specialist stem cell biotechnology company, Stemnovate Limited, in Cambridge, UK.

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Our recombinant human TGF-β1 PLUS protein requires careful handling due to its poor solubility in physiological solutions. Please follow the handling guidance for lyophilized cytokines below to minimize loss of protein due to precipitation or adsorption to plastic. We advise storing the recombinant protein at very low pH before dilution in cell culture media or final working solutions. Low pH will also assist in maintaining the correct disulfide structure of the protein by minimizing disulfide bond exchange reactions.

  • Resuspension in physiological buffers may cause precipitation of stock solutions, hence we recommend dissolving our lyophilized cytokines in 10 mM HCl (1:1000 dilution of concentrated HCl) while keeping the protein concentration at 50 µg/ml or above, in order to avoid loss by adsorption to plasticware.
  • To ensure you recover all of the protein, let the sample sit for a few minutes with the solubilization buffer at room temperature and pipette gently up and down (avoid foaming).
  • Rinse the tube with some more 10 mM HCl and pool with the rest.
  • The protein is tolerant of some freeze and thaw cycles, but as always with proteins, it is better to aliquot and store frozen.
  • Our proteins are supplied carrier protein-free. If compatible with your work, add carrier protein of your choice such as BSA, HSA or gelatin to further minimize loss by adsorption.
  • Store in -80°C for long-term storage, -20°C for short-term storage.

We 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.

Recovery: protein quantitation

Result: UV spectrum shows full recovery of protein following aliquoting and lyophilization.

Absorbance at 280 nm: average 0.22
Recovered concentration: 0.22 cm-1 x 10 / 2.21 cm-1 mg ml-1 = 1.00 mg / ml
Recovery: 100%

Concentration was calculated using extinction coefficient at 280 nm. Example data from Qk010 batch #012.

Product Documents

Certificate of Analysis

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