Recombinant Human Wnt-3a GMP Protein, CF GMP
R&D Systems, part of Bio-Techne | Catalog # 5036-GMP
Key Product Details
Product Specifications
Source
Chinese Hamster Ovary cell line, CHO-derived human Wnt-3a protein
Ser19-Lys352
Manufactured and tested under cGMP guidelines.
Ser19-Lys352
Manufactured and tested under cGMP guidelines.
Purity
>75%, by SDS-PAGE with silver staining, under reducing conditions.
Endotoxin Level
<0.10 EU per 1 μg of the protein by the LAL method.
N-terminal Sequence Analysis
Ser19-Tyr-Pro-Ile-Trp-Trp-Ser-Leu-Ala-Val
Predicted Molecular Mass
37.4 kDa
SDS-PAGE
40 kDa, reducing conditions
Activity
Measured by its ability to induce alkaline phosphatase production by MC3T3-E1 mouse preosteoblast cells.
The ED50 for this effect is 5-25 ng/mL.
Measured by its ability to induce Topflash reporter activity in HEK293T human embryonic kidney cells.
The ED50 for this effect is <500 ng/mL. Protein concentrations should be titrated based on cell type and if appropriate, passage number of the cell line.
Optimal concentrations should be determined by each laboratory for each application.
The ED50 for this effect is 5-25 ng/mL.
Measured by its ability to induce Topflash reporter activity in HEK293T human embryonic kidney cells.
The ED50 for this effect is <500 ng/mL. Protein concentrations should be titrated based on cell type and if appropriate, passage number of the cell line.
Optimal concentrations should be determined by each laboratory for each application.
Host Cell Protein
< 40 ng per µg of protein when tested by ELISA.
Mycoplasma
Negative when tested in a ribosomal RNA hybridization assay.
Scientific Data Images for Recombinant Human Wnt-3a GMP Protein, CF
Recombinant Human Wnt-3a GMP Protein Bioactivity
GMP-grade Recombinant Human Wnt-3a (Catalog # 5036-GMP) induces alkaline phosphatase production by the MC3T3‑E1 mouse preosteoblast cell line. The ED50 for this effect is 5-25 ng/mL. Three independent lots were tested for activity and plotted on the same graph to show lot-to-lot consistency of GMP Wnt-3a.
Recombinant Human Wnt-3a GMP Protein Bioactivity
GMP-grade Recombinant Human Wnt-3a (Catalog # 5036-GMP) induces beta-catenin-responsive transcriptional activation in a Topflash Luciferase assay using the HEK293T human embryonic kidney cell line. The ED50 for this effect is <500 ng/mL. Three independent lots were tested for activity and plotted on the same graph to show lot-to-lot consistency of GMP Wnt-3a.Formulation, Preparation and Storage
5036-GMP
| Formulation | Lyophilized from a 0.2 μm filtered solution in PBS, EDTA and CHAPS. |
| Reconstitution | Reconstitute at 200 μg/mL in PBS. |
| Shipping | The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below. |
| Stability & Storage | Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
|
Background: Wnt-3a
Wnt-3a is one of 19 vertebrate members of the Wingless-type MMTV integration site (Wnt) family of highly conserved cysteine-rich secreted glycoproteins important for normal developmental processes (1). Wnts bind to the cell surface Frizzled family receptors in conjunction with low-density lipoprotein receptor-related protein family receptors (LRP5 or 6) resulting in the stabilization of intracellular beta-catenin levels (2). As intracellular beta-catenin levels rise, beta-catenin binds to TCF/LEF transcription factors leading to expression of Wnt target genes (3). Endo-IWR 1 (Catalog # 3532, # PSM1324) is a cell-permeant small molecule inhibitor of Axin turnover that suppresses Wnt signal transduction by stabilizing the beta-catenin destruction complex (4). Wnt-3a is a 44 kDa secreted hydrophobic glycoprotein containing a conserved pattern of 24 cysteine residues (5). Wnt-3a has two N-linked glycosylation sites (Asn 87, Asn 298), and Ser 209 is modified with palmitoleic acid (6). Glycosylation and acylation are essential for efficient Wnt secretion and biological activity, respectively (6, 7). Human Wnt-3a shares 96% amino acid (aa) identity with mouse mouse, bovine and canine Wnt-3a, and 89%, 86% and 84% aa identity with chicken, Xenopus and zebrafish Wnt-3a, respectively. It also shares 87% aa identity with Wnt3. During embryonic development, Wnt-3a is necessary for proper development of the hippocampus, anterior-posterior patterning, somite development, and tailbud formation (9-12). Wnt-3a also promotes self-renewal of hematopoietic stem cells, neural stem cells, and embryonic stem cells (13, 14).
References
- Willert, K. and Nusse, R. (2012) Cold Spring Harb. Perspect. Biol. 4:a007864.
- MacDonald, B.T. and X. He (2012) Cold Spring Harb. Perspect. Biol. 4:a007880.
- Korinek, V. et al. (1997) Science 275:1784.
- Chen, B. et al. (2009) Nat. Chem. Biol. 5:100.
- Smolich, B.D. et al. (1993) Mol. Biol. Cell 4:1267.
- Takada, R. et al. (2006) Dev. Cell 11:791.
- Komekado, H. (2007) Genes Cells 12:521.
- Dunty Jr. W. C. et al. (2008) Development 135:85.
- Ikeya, M. and S. Takada (2001) Mech. Dev. 103:27.
- Lee, S. M. et al. (2000) Development 127:457.
- Takada, S. et al. (1994) Genes Dev. 8:174.
- Willert, K. et al. (2003) Nature 423:6938.
- Kalani, M.Y. et al. (2008) Proc. Natl. Acad. Sci. USA 105:16970.
- Ten Berge, D. et al. (2011) Nat. Cell Biol. 13:1070.
Long Name
Wingless-type MMTV Integration Site Family, Member 3a
Alternate Names
Wnt3a
Gene Symbol
WNT3A
UniProt
Additional Wnt-3a Products
Product Documents for Recombinant Human Wnt-3a GMP Protein, CF
Manufacturing Specifications
GMP ProteinsR&D Systems, a Bio-Techne Brand's GMP proteins are produced according to relevant sections of the following documents: USP Chapter 1043, Ancillary Materials for Cell, Gene and Tissue-Engineered Products and Eu. Ph. 5.2.12, Raw Materials of Biological Origin for the Production of Cell-based and Gene Therapy Medicinal Products.
R&D Systems' quality focus includes:
- Manufactured and tested under an ISO 9001:2015 and ISO 13485:2016 certified quality system
- Documented processes and QA control of documentation and process changes
- Personnel training programs
- Raw material testing and vendor qualification/monitoring
- Fully validated equipment, processes and test methods
- Equipment calibration schedules using a computerized calibration program
- Facility maintenance, safety programs and pest control
- Material review process for variances
- Monitoring of stability over product shelf-life
- N-terminal amino acid analysis, SDS-PAGE analysis, and endotoxin level (as determined by LAL assay) performed on each bulk QC lot, not on individual bottlings of each QC lot
- Post-bottling lot-specific bioassay results (compliance with an established range) and results of microbial testing according to USP
- Host Cell Protein testing performed by ELISA
- Mycoplasma testing by ribosomal RNA hybridization assay
Additional testing and documentation requested by the customer can be arranged at an additional cost.
Production records and facilities are available for examination by appropriate personnel on-site at R&D Systems in Minneapolis, Minnesota USA.
R&D Systems sells GMP grade products for preclinical or clinical ex vivo use. They are not for in vivo use. Please read the following End User Terms prior to using this product.
Product Specific Notices for Recombinant Human Wnt-3a GMP Protein, CF
END USER TERMS OF USE OF PRODUCT
The following terms are offered to you upon your acceptance of these End User Terms of Use of Product. By using this product, you indicate your acknowledgment and agreement to these End User Terms of Use of Product. If you do not agree to be bound by and comply with all of the provisions of these End User Terms of Use of Product, you should contact your supplier of the product and make arrangements to return the product.
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The End User is aware that R&D Systems, Inc. sells GMP products for preclinical or clinical ex vivo use and not for in vivo use. The End User further agrees, as a condition of the sale of R&D Systems' GMP products that: a) the End User will not use this GMP Product in any procedure wherein the product may be directly or indirectly administered to humans, unless the End User has obtained, or prior to their use will have obtained, an Investigational New Drug (IND) exemption from the FDA and will use the product only in accordance with the protocols of such IND and of the Institutional Review Board overseeing the proposed research, or b) the End User will use the products outside of the United States in accordance with the protocols of research approved by the Institutional Review Board or authorized ethics committee and regulatory agencies to which the End User is subject to in their territory.
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For preclinical, or clinical ex vivo use
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