Recombinant SARS-CoV-2 Spike RBD His-tag Protein, CF

Name: Recombinant SARS-CoV-2 Spike RBD His-tag Protein, CF
Brand: R&D Systems
SKU: 10500-CV

R&D Systems, Inc. a Bio-Techne Brand

Catalog # 10500-CV

Citations (1)

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HEK293 Expressed, High ACE-2 Binding

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Key Product Details

Source:	HEK293
Accession #:	YP_009724390.1
Applications:	Bioactivity

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Product Specifications for Recombinant SARS-CoV-2 Spike RBD His-tag Protein, CF

Source	Human embryonic kidney cell, HEK293-derived sars-cov-2 Spike RBD protein Arg319-Phe541, with a C-terminal 6-His tag
Purity	>95%, by SDS-PAGE visualized with Silver Staining and quantitative densitometry by Coomassie® Blue Staining.
Endotoxin level	<0.10 EU per 1 μg of the protein by the LAL method.
N-terminal sequence Analysis	Arg319
Predicted Molecular Mass	26 kDa
SDS-PAGE	28-38 kDa, under reducing conditions
Activity	Measured by its binding ability in a functional ELISA with Recombinant Human ACE-2 His-tag (Catalog # 933-ZN).

Scientific Data Examples for Recombinant SARS-CoV-2 Spike RBD His-tag Protein, CF

	Recombinant SARS-CoV-2 Spike RBD His-tag Protein Bioactivity Recombinant SARS-CoV-2 Spike RBD His-tag Protein (HEK293 Expressed) (10500-CV) binds Recombinant Human ACE-2 His-tag (933-ZN) in a functional ELISA.
	Recombinant SARS-CoV-2 Spike RBD His-tag Protein SDS-PAGE 2 μg/lane of Recombinant SARS-CoV-2 Spike RBD His-tag Protein (HEK293 Expressed) (10500-CV) was resolved with SDS-PAGE under reducing (R) and non-reducing (NR) conditions and visualized by Coomassie® Blue staining, showing bands at 28-38 kDa.
	Binding of ACE-2 to SARS-CoV-2 Spike RBD by surface plasmon resonance (SPR). Recombinant SARS-CoV-2 Spike RBD His-tag Protein (Catalog # 10500-CV) was immobilized on a Biacore Sensor Chip CM5, and binding to recombinant human ACE-2 (Catalog # 933-ZN) was measured at a concentration range between 0.37 nM and 93.5 nM. The double-referenced sensorgram was fit to a 1:1 binding model to determine the binding kinetics and affinity, with an affinity constant of KD=2.149 nM. (Biacore T200).
	R&D Systems Spike RBD protein beats the competition. Recombinant SARS-CoV-2 Spike RBD (10500-CV) binds recombinant human ACE-2 (10544-ZN) in a functional ELISA. The binding activity is approximately 3-fold greater than a top competitor’s Spike RBD protein.
	SDS-PAGE of 1 µg of R&D Systems Recombinant SARS-CoV-2 Spike RBD (10500-CV) or a competitor’s Spike RBD protein were run under reducing or non-reducing conditions and visualized by silver staining. The R&D Systems Spike protein runs as a sharper, more definitive band than the competitor’s Spike RBD protein.

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.

10500-CV

Formulation	Lyophilized from a 0.2 μm filtered solution in PBS with Trehalose.
Reconstitution	Reconstitute at 500 μg/mL in PBS.
Shipping	The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below.
Stability & Storage:	Use a manual defrost freezer and avoid repeated freeze-thaw cycles. 12 months from date of receipt, -20 to -70 °C as supplied. 1 month, 2 to 8 °C under sterile conditions after reconstitution. 3 months, -20 to -70 °C under sterile conditions after reconstitution.

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Background: Spike RBD

SARS-CoV-2, which causes the global pandemic coronavirus disease 2019 (Covid-19), belongs to a family of viruses known as coronaviruses that are commonly comprised of four structural proteins: Spike protein(S), Envelope protein (E), Membrane protein (M), and Nucleocapsid protein (N) (1). SARS-CoV-2 Spike Protein (S Protein) is a glycoprotein that mediates membrane fusion and viral entry. The S protein is homotrimeric, with each ~180-kDa monomer consisting of two subunits, S1 and S2 (2). In SARS-CoV-2, as with most coronaviruses, proteolytic cleavage of the S protein into two distinct peptides, S1 and S2 subunits, is required for activation. The S1 subunit is focused on attachment of the protein to the host receptor while the S2 subunit is involved with cell fusion (3-5). Based on structural biology studies, the receptor binding domain (RBD), located in the C-terminal region of S1, can be oriented either in the up/standing or down/lying state (6). The standing state is associated with higher pathogenicity and both SARS-CoV-1 and MERS can access this state due to the flexibility in their respective RBDs. A similar two-state structure and flexibility is found in the SARS-CoV-2 RBD (7). Based on amino acid (aa) sequence homology, the SARS-CoV-2 S1 subunit RBD has 73% identity with the RBD of the SARS-CoV-1 S1 RBD, but only 22% homology with the MERS S1 RBD. The low aa sequence homology is consistent with the finding that SARS and MERS bind different cellular receptors (8). The S Protein of the SARS-CoV-2 virus, like the SARS-CoV-1 counterpart, binds Angiotensin-Converting Enzyme 2 (ACE2), but with much higher affinity and faster binding kinetics (9). Before binding to the ACE2 receptor, structural analysis of the S1 trimer shows that only one of the three RBD domains in the trimeric structure is in the "up" conformation. This is an unstable and transient state that passes between trimeric subunits but is nevertheless an exposed state to be targeted for neutralizing antibody therapy (10). Polyclonal antibodies to the RBD of the SARS-CoV-2 protein have been shown to inhibit interaction with the ACE2 receptor, confirming RBD as an attractive target for vaccinations or antiviral therapy (11). There is also promising work showing that the RBD may be used to detect presence of neutralizing antibodies present in a patient's bloodstream, consistent with developed immunity after exposure to the SARS-CoV-2 virus (12). Lastly, it has been demonstrated the S Protein can invade host cells through the CD147/EMMPRIN receptor and mediate membrane fusion (13, 14).

Item	Value
References	Wu, F. et al. (2020) Nature 579:265. Tortorici, M.A. and D. Veesler (2019). Adv. Virus Res. 105:93. Bosch, B.J. et al. (2003) J. Virol. 77:8801. Belouzard, S. et al. (2009) Proc. Natl. Acad. Sci. 106:5871. Millet, J.K. and G. R. Whittaker (2015) Virus Res. 202:120. Yuan, Y. et al. (2017) Nat. Commun. 8:15092. Walls, A.C. et al. (2010) Cell 180:281. Jiang, S. et al. (2020) Trends. Immunol. https://doi.org/10.1016/j.it.2020.03.007. Ortega, J.T. et al. (2020) EXCLI J. 19:410. Wrapp, D. et al. (2020) Science 367:1260. Tai, W. et al. (2020) Cell. Mol. Immunol. https://doi.org/10.1016/j.it.2020.03.007. Okba, N. M. A. et al. (2020). Emerg. Infect. Dis. https://doi.org/10.3201/eid2607.200841. Wang, X. et al. (2020) https://doi.org/10.1038/s41423-020-0424-9. Wang, K. et al. (2020) bioRxiv https://www.biorxiv.org/content/10.1101/2020.03.14.988345v1.
Long Name	Spike Receptor Binding Domain
Entrez Gene IDs	3200426 (HCoV-HKU1); 14254594 (MERS-CoV); 1489668 (SARS-CoV); 43740568 (SARS-CoV-2)
Alternate Names	Spike RBD

Citations for Recombinant SARS-CoV-2 Spike RBD His-tag Protein, CF (1)

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Citations are publications that use Bio-Techne products. Selected citations for Recombinant SARS-CoV-2 Spike RBD His-tag Protein, CF include:

Species: Human
Sample Types: Serum
Applications: ELISA Capture

JH Chang et al. (2022), Humoral Immunogenicity and Reactogenicity of the Standard ChAdOx1 nCoV-19 Vaccination in Taiwan Vaccines, 10(2).
PMID: 35214770

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