Recombinant Human Wnt-3a GMP Protein, CFGMP

Name: Recombinant Human Wnt-3a GMP Protein, CF
Brand: R&D Systems
SKU: 5036-GMP

Catalog # 5036-GMP

Citations (1)

Product Datasheet / COA / SDS

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Key Product Details

Source:	CHO
Accession #:	NP_149122
Applications:	Bioactivity

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Product Specifications

Source	Chinese Hamster Ovary cell line, CHO-derived human Wnt-3a protein 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.

Recombinant Human Wnt-3a GMP Protein, CF Scientific Data Examples

	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

Carrier Free

What does CF mean?

CF stands for Carrier Free (CF). We typically add Bovine Serum Albumin (BSA) as a carrier protein to our recombinant proteins. Adding a carrier protein enhances protein stability, increases shelf-life, and allows the recombinant protein to be stored at a more dilute concentration. The carrier free version does not contain BSA.

What formulation is right for me?

In general, we advise purchasing the recombinant protein with BSA for use in cell or tissue culture, or as an ELISA standard. In contrast, the carrier free protein is recommended for applications, in which the presence of BSA could interfere.

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. A minimum of 6 months when stored at ≤ -20 °C as supplied. Refer to lot specific COA for the Use by Date. 1 month, 2 to 8 °C under sterile conditions after reconstitution. 3 months, ≤ -20 °C under sterile conditions after reconstitution.

Reconstitution Calculator

The reconstitution calculator allows you to quickly calculate the volume of a reagent to reconstitute your vial. Simply enter the mass of reagent and the target concentration and the calculator will determine the rest.

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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).

Item	Value
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
Entrez Gene IDs	89780 (Human); 22416 (Mouse)
Alternate Names	MGC119418, MGC119419, MGC119420, Wnt-3a, Wnt3a, protein Wnt-3a, wingless-type MMTV integration site family, member 3A

Manufacturing Specifications

GMP Proteins
R&D Systems, a Bio-Techne Brand's GMP proteins are produced according to relevant sections of the following documents: WHO TRS, No. 822, 1992 Annex 1, Good Manufacturing Practices for Biological Products; USP Chapter 1043, Ancillary Materials for Cell, Gene and Tissue-Engineered Products and USP Chapter 92, Growth Factors and Cytokines Used in Cell Therapy Manufacturing.

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

R&D Systems strives to provide our customers with the analytical characteristics of each product so that customers may determine whether our products are appropriate for their research. The Certificate of Analysis provided contains the following lot specific information:

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

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.

We suggest you print and retain a copy of these End User Terms of Use of Product for your records.

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.

R&D Systems, Inc. has the right, at its sole discretion, to modify, add or remove any terms or conditions of these End User Terms of Use without notice or liability to you. Any changes to these End User Terms of Use are effective immediately following the printing of such changes on this product insert. The most recent version of these End User Terms of Use of Product may be found at: RnDSystems.com/Legal .

You agree to review these End User Terms of Use of Product to ensure any subsequent use by you of R&D Systems' GMP Products following changes to these End User Terms of Use of Product constitutes your acceptance of all such changes.

TERMS AND CONDITIONS

The following limitation applies to R&D Systems' warranty and liability for damages: All products are warranted to meet R&D Systems' published specifications when used under normal laboratory conditions.

R&D SYSTEMS DOES NOT MAKE ANY OTHER WARRANTY OR REPRESENTATION WHATSOEVER, WHETHER EXPRESS OR IMPLIED, WITH RESPECT TO ITS PRODUCTS. IN PARTICULAR, R&D SYSTEMS DOES NOT MAKE ANY WARRANTY OF SUITABILITY, NONINFRINGEMENT, MERCHANTABILITY OR FITNESS FOR ANY PARTICULAR PURPOSE.

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Citations for Recombinant Human Wnt-3a GMP Protein, CF (1)

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Citations are publications that use Bio-Techne products. Selected citations for Recombinant Human Wnt-3a GMP Protein, CF include:

Species: Human
Sample Types: Whole Cells

SG Cho (2017), APC downregulated 1 inhibits breast cancer cell invasion by inhibiting the canonical WNT signaling pathway Oncol Lett, 14(4):4845-4852.
PMID: 29085490

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