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Recombinant human R-spondin 1 protein

QK006

Brand: Qkine

R-spondin 1 protein (RSPO1) is the prototypic member of the R-spondin family and is used to potentiate Wnt signaling in many organoid culture systems including intestinal and tumor (cancer) organoid culture. R-spondin 1 is also required for hematopoietic stem cell specification and cancer cell migration and survival.

Recombinant human R-spondin 1 protein (RSPO1) protein is the bioactive domain of human R-spondin 1 comprising the two cysteine-rich furin-like domains of R-spondin 1. Those are necessary and sufficient for Wnt signaling potentiation and are the essential domains for activity in stem cell and organoid culture.

This protein has a molecular weight of 13 kDa and is animal origin-free (AOF), carrier protein-free, and tag-free to ensure its purity with exceptional lot-to-lot consistency. Qk006 is suitable to replace R-spondin conditioned media for improved reproducibility in chemically defined organoid culture media.

Currency: 

Product name Catalog number Pack size Price Price (USD) Price (GBP) Price (EUR)
Recombinant human R-spondin 1 protein, 25 µg QK006-0025 25 µg (select above) $ 210.00 £ 155.00 € 182.00
Recombinant human R-spondin 1 protein, 50 µg QK006-0050 50 µg (select above) $ 290.00 £ 210.00 € 246.00
Recombinant human R-spondin 1 protein, 100 µg QK006-0100 100 µg (select above) $ 465.00 £ 340.00 € 398.00
Recombinant human R-spondin 1 protein, 500 µg QK006-0500 500 µg (select above) $ 1,650.00 £ 1,200.00 € 1,402.00
Recombinant human R-spondin 1 protein, 1000 µg QK006-1000 1000 µg (select above) $ 2,600.00 £ 1,900.00 € 2,220.00

Note: prices shown do not include shipping and handling charges.

Qkine company name and logo are the property of Qkine Ltd. UK.

Alternative protein names
Roof plate-specific spondin 1, Rspo1, R-spondin 1, Cristin 3
Species reactivity

Human

Species similarity:
Porcine – 98%
Mouse – 96%
Rat – 96%
Bovine – 94%

Alternative:

R-spondin 1 LR5 protein (Qk031): a specialized form of R-spondin 1 that specifically binds to the LGR5 receptor, developed in the lab of Marc de la Roche (University of Cambridge).

Frequently used together with:

Summary

  • High purity recombinant human R-spondin 1 protein (Uniprot: Q2MKA7)
  • >98%, by SDS-PAGE quantitative densitometry
  • 13 kDa
  • Expressed in E. coli
  • Animal origin-free (AOF) and carrier protein-free.
  • Manufactured in Qkine's Cambridge, UK laboratories
  • Lyophilized from acetonitrile, TFA
  • Resuspend in 10mM HCl at >100 µg/ml (provided with protein and free of charge), prepare single use aliquots, add carrier protein if desired and store frozen at -20°C or -80°C
Handling and Storage FAQ

Featured applications

  • Pancreatic tumor organoid culture
  • Maintenance of urine-derived stem cells

Bioactivity

Human R-spondin 1 Qk006 protein bioactivity lot #012

R-spondin 1 (Qk006) activity is determined using the Wnt-responsive firefly luciferase reporter assay as it enhances Wnt-ß catenin signaling in HEK239T cells. HEK293T cells transfected with reporter TOP-FLASH are treated in triplicate with increasing concentration of R-spondin 1 (diluted in DMEM with 0.5 % of FCS), in the presence of Wnt-conditioned media (1:8 dilution). Cells are grown overnight, and luciferase activity is measured and normalized. EC50 = 5.8 nM (75.1 ng/mL). Data from Qk006 lot #104277.

Purity

Human R-spondin 1 Qk006 protein purity SDS-PAGE lot #010

R-spondin 1 migrates as a single band at 16 kDa in non-reducing (NR) and 13 kDa in reducing (R) conditions. 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.  Data from Qk006 lot #010.

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%

R-spondin 1 (Qk006) has be tested at 100 ng/ml – 500 ng/ml, we would recommend using 100 ng/ml – 1 µg/ml for initial testing and optimisation with culture. It is highly recommended to do a dose response to optimize concentration due to varying culture conditions across different labs.

Qkine FGF-4 bioactivity is comparable to an alternative supplier protein

Quantitative luciferase assay with Qkine FGF-4 (Qk004, green) and alternative supplier FGF-4 (Supplier B, black). Cells were treated in triplicate with a serial dilution of FGF-4 for 3 hours. Firefly luciferase activity was measured and normalized to control Renilla luciferase activity.

Protein background

R-spondin 1 (RSPO-1) is a secreted protein of the RSPO family that regulates the canonical Wnt signaling pathway [1,2]. The Wnt pathway is a highly conserved signaling pathway that is essential for various cellular processes, including cell proliferation, differentiation, and migration. This growth factor plays a crucial role in embryonic development and patterning, particularly in the development of the gastrointestinal tract, neural tube, skin, and limbs [3]. It is involved in the maintenance of stem cells and can enhance the self-renewal capacity of stem cells, promoting tissue regeneration and repair [4]. It is also involved in the sex determination and the development and maintenance of the female sex characteristics in particular the differentiation and maintenance of the ovarian tissues [5].

RSPO1 proteins are generally characterized by a core domain known as the thrombospondin type 1 (TSP1) domain, which is responsible for their signaling activity. This protein is comprised of 263 amino acids and structured as two cysteine-rich, furin-like domains [3,6]. It acts as an agonist (activator) of the Wnt signaling pathway by binding to the leucine-rich repeat-containing G-protein coupled receptors (LGR) 4-6, leading to the stabilization of β-catenin, a key component of the pathway [7].

In cell culture, this recombinant protein is used extensively for the maintenance of self-renewal and expansion of stem cells. It is also used for the culture of intestinal, hematopoietic, and epithelial stem cells [2,7,8]. Also, the culture of stem cell-derived organoids such as intestinal, pancreas liver, colon, and stomach organoids benefit from the inclusion of  RSPO1 in the media [7,9,10].

Dysregulation of the Wnt signaling pathway, including the overexpression of R-spondin 1, has been associated with tumor growth and progression in various cancers such as colon and ovarian cancer [11,12]. Research is ongoing on the role of R-spondin 1 in various biological processes and its potential as a therapeutic target for cancer treatment. Given its role in stem cell maintenance and tissue regeneration,  RSPO1 and related proteins have also been explored as potential therapeutic targets for regenerative medicine.

Background references

  1. K.-A. Kim et al., ‘R-Spondin family members regulate the Wnt pathway by a common mechanism’, Mol. Biol. Cell, vol. 19, no. 6, pp. 2588–2596, Jun. 2008, doi: 10.1091/mbc.e08-02-0187.
  2. W. de Lau, W. C. Peng, P. Gros, and H. Clevers, ‘The R-spondin/Lgr5/Rnf43 module: regulator of Wnt signal strength’, Genes Dev., vol. 28, no. 4, pp. 305–316, Feb. 2014, doi: 10.1101/gad.235473.113.
  3. A. Glinka et al., ‘LGR4 and LGR5 are R-spondin receptors mediating Wnt/β-catenin and Wnt/PCP signalling’, EMBO Rep., vol. 12, no. 10, pp. 1055–1061, Sep. 2011, doi: 10.1038/embor.2011.175.
  4. K. S. Yan et al., ‘Non-equivalence of Wnt and R-spondin ligands during Lgr5+ intestinal stem-cell self-renewal’, Nature, vol. 545, no. 7653, Art. no. 7653, May 2017, doi: 10.1038/nature22313.
  5. K. Tomizuka et al., ‘R-spondin1 plays an essential role in ovarian development through positively regulating Wnt-4 signaling’, Hum. Mol. Genet., vol. 17, no. 9, pp. 1278–1291, May 2008, doi: 10.1093/hmg/ddn036.
  6. W. B. de Lau, B. Snel, and H. C. Clevers, ‘The R-spondin protein family’, Genome Biol., vol. 13, no. 3, p. 242, Mar. 2012, doi: 10.1186/gb-2012-13-3-242.
  7. W. de Lau et al., ‘Lgr5 homologues associate with Wnt receptors and mediate R-spondin signalling’, Nature, vol. 476, no. 7360, Art. no. 7360, Aug. 2011, doi: 10.1038/nature10337.
  8. J. R. Genthe and W. K. Clements, ‘R-spondin 1 is required for specification of hematopoietic stem cells through Wnt16 and Vegfa signaling pathways’, Development, vol. 144, no. 4, pp. 590–600, Feb. 2017, doi: 10.1242/dev.139956.
  9. L. Broutier et al., ‘Culture and establishment of self-renewing human and mouse adult liver and pancreas 3D organoids and their genetic manipulation’, Nat. Protoc., vol. 11, no. 9, Art. no. 9, Sep. 2016, doi: 10.1038/nprot.2016.097.
  10. L. Broutier et al., ‘Human primary liver cancer–derived organoid cultures for disease modeling and drug screening’, Nat. Med., vol. 23, no. 12, Art. no. 12, Dec. 2017, doi: 10.1038/nm.4438.
  11. S. Seshagiri et al., ‘Recurrent R-spondin fusions in colon cancer’, Nature, vol. 488, no. 7413, Art. no. 7413, Aug. 2012, doi: 10.1038/nature11282.
  12. E. J. ter Steege and E. R. M. Bakker, ‘The role of R-spondin proteins in cancer biology’, Oncogene, vol. 40, no. 47, Art. no. 47, Nov. 2021, doi: 10.1038/s41388-021-02059-y.

Publications using recombinant human R-spondin 1 protein (Qk006)

Human epidermis organotypic cultures, a reproducible system recapitulating the epidermis in vitro
In Experimental Dermatology on 28 April 2023 by Agarwal, R. et al.
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 Establishment of Patient-Derived Organoids Using Ascitic or Pleural Fluid from Cancer Patients

In Cancer Research and Treatment on 12 June 2023 by Choi, W et al.
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The effect of extracellular matrix on the precision medicine utility of pancreatic cancer patient–derived organoids.
In JCI Insight on 5 December 2023 by Lumibao, J.C., Okhovat, S.R, et al.
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 PRMT1 promotes pancreatic cancer development and resistance to chemotherapy

In Cells Reports Medicine on 8 March 2024 by Ku, B., Eisenbarth, D., et al.
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