Recombinant SARSCoV-2 B.1.1.529 RBD Alexa Fluor® 488 Protein

Name: Recombinant SARSCoV-2 B.1.1.529 RBD Alexa Fluor® 488 Protein
Brand: R&D Systems
SKU: AFG11056

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

Catalog # AFG11056

Product Datasheet / COA / SDS

Omicron Variant His-tag

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Key Product Details

Source:	HEK293
Accession #:	YP_009724390.1
Structure / Form:	Labeled with Alexa Fluor® 488 via amines Excitation Wavelength: 488 nm Emission Wavelength: 515-545 nm
Applications:	Bioactivity

View Full Technical Details for Recombinant SARSCoV-2 B.1.1.529 RBD Alexa Fluor® 488 Protein

Product Specifications for Recombinant SARSCoV-2 B.1.1.529 RBD Alexa Fluor® 488 Protein

Source	Human embryonic kidney cell, HEK293-derived sars-cov-2 Spike RBD protein Arg319-Phe541 (Gly339Asp, Ser371Leu, Ser373Pro, Ser375Phe, Lys417Asn, Asn440Lys, Gly446Ser, Ser477Asn, Thr478Lys, Glu484Ala, Gln493Arg, Gly496Ser, Gln498Arg, Asn501Tyr, Tyr505His), 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	<1.0 EU per 1 μg of the protein by the LAL method.
N-terminal sequence Analysis	Arg319
Predicted Molecular Mass	26 kDa
SDS-PAGE	33-39 kDa, under reducing conditions.
Activity	Measured by flow cytometry for its ability to bind HEK293 human embryonic kidney cells transfected with human ACE-2 at 0.250-1.00 µg/mL (100 µL/well). Please Note: Optimal dilutions should be determined by each laboratory for each application.

Scientific Data Examples for Recombinant SARSCoV-2 B.1.1.529 RBD Alexa Fluor® 488 Protein

	Detection of ACE-2 on HEK293 Transfectants with Recombinant SARS-CoV-2 B.1.1.529 Spike RBD His-tag Alexa Fluor® 488 Protein by Flow Cytometry. HEK293 human embryonic kidney cells transfected with human ACE-2 were stained with (A) Recombinant SARS-CoV-2 B.1.1.529 Spike RBD His-tag Alexa Fluor® 488 (Catalog # AFG11056, filled histogram) or (B) unstained (open histogram).
	Recombinant SARS-CoV-2 B.1.1.529 Spike RBD His-tag Alexa Fluor® 488 Protein SDS-PAGE. 2 μg/lane of Recombinant SARS-CoV-2 B.1.1.529 Spike RBD His-tag Alexa Fluor® 488 Protein (Catalog # AFG11056) was resolved with SDS-PAGE under reducing (R) and non-reducing (NR) conditions and visualized by Coomassie® Blue staining, showing bands at 33-39 kDa.

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.

AFG11056

Formulation	Supplied as a 0.2 μm filtered solution in PBS with BSA as a carrier protein.
Shipping	The product is shipped with dry ice or equivalent. Upon receipt, store it immediately at the temperature recommended below.
Stability & Storage:	Protect from light. Use a manual defrost freezer and avoid repeated freeze-thaw cycles. 6 months from date of receipt, -20 to -70 °C as supplied. 1 month, 2 to 8 °C under sterile conditions after opening. 3 months, -20 to -70 °C under sterile conditions after opening.

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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 also include MERS‑CoV and SARS-CoV-1. Coronaviruses are commonly comprised of four structural proteins: Spike protein (S), Envelope protein (E), Membrane protein (M) and Nucleocapsid protein (N) (1). The SARS-CoV-2 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 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). A receptor binding domain (RBD) in the C-terminus of the S1 subunit has been identified and the RBD of SARS-CoV-2 shares 73% amino acid (aa) identity with the RBD of the SARS-CoV-1, but only 22% aa identity with the RBD of MERS‑CoV (6, 7). The low aa sequence homology is consistent with the finding that SARS and MERS‑CoV bind different cellular receptors (8). The RBD of SARS-CoV-2 binds a metallopeptidase, angiotensin-converting enzyme 2 (ACE-2), similar to SARS-CoV-1, but with much higher affinity and faster binding kinetics (9). Before binding to the ACE-2 receptor, structural analysis of the S1 trimer shows that only one of the three RBD domains 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 ACE-2 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 (12). Several emerging SARS-CoV-2 genomes have been identified including the Omicron, or B.1.1.529, variant. First identified in November 2021 in South Africa, the Omicron variant quickly became the predominant SARS-CoV-2 variant and is considered a variant of concern (VOC). The Omicron variant contains 15 mutations in RBD domain that potentially affect viral fitness and transmissibility. The majority of the mutations are involved in ACE-2 binding and Omicron binds ACE-2 with greater affinity, potentially explaining its increased transmissibility (13, 14). Several of these mutations are also identified in facilitating immune escape and reducing neutralization activity to several monoclonal antibodies (13). Additionally, a series of new mutations are present in the RBD which have unknown impacts on receptor binding or antibody neutralization.

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. Li, W. et al. (2003) Nature 426:450. Wong, S.K. et al. (2004) J. Biol. Chem. 279:3197. 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.< /li> Wrapp, D. et al. (2020) Science 367:1260. Tai, W. et al. (2020) Cell. Mol. Immunol. 17:613. Okba, N. M. A. et al. (2020). Emerg. Infect. Dis. https://doi.org/10.3201/eid2607.200841. Shah, M. and H.G. Woo (2021) bioRxiv https://doi.org/10.1101/2021.12.04.471200. Lupala, C.S. et al. (2021) bioRxiv https://doi.org/10.1101/2021.12.10.472102.
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

Product Specific Notices for Recombinant SARSCoV-2 B.1.1.529 RBD Alexa Fluor® 488 Protein

This product is provided under an agreement between Life Technologies Corporation and R&D Systems, Inc, and the manufacture, use, sale or import of this product is subject to one or more US patents and corresponding non-US equivalents, owned by Life Technologies Corporation and its affiliates. The purchase of this product conveys to the buyer the non-transferable right to use the purchased amount of the product and components of the product only in research conducted by the buyer (whether the buyer is an academic or for-profit entity). The sale of this product is expressly conditioned on the buyer not using the product or its components (1) in manufacturing; (2) to provide a service, information, or data to an unaffiliated third party for payment; (3) for therapeutic, diagnostic or prophylactic purposes; (4) to resell, sell, or otherwise transfer this product or its components to any third party, or for any other commercial purpose. Life Technologies Corporation will not assert a claim against the buyer of the infringement of the above patents based on the manufacture, use or sale of a commercial product developed in research by the buyer in which this product or its components was employed, provided that neither this product nor any of its components was used in the manufacture of such product. For information on purchasing a license to this product for purposes other than research, contact Life Technologies Corporation, Cell Analysis Business Unit, Business Development, 29851 Willow Creek Road, Eugene, OR 97402, Tel: (541) 465-8300. Fax: (541) 335-0354.
This product is provided under an agreement between Life Technologies Corporation and R&D Systems, Inc, and the manufacture, use, sale or import of this product is subject to one or more US patents and corresponding non-US equivalents, owned by Life Technologies Corporation and its affiliates. The purchase of this product conveys to the buyer the non-transferable right to use the purchased amount of the product and components of the product only in research conducted by the buyer (whether the buyer is an academic or for-profit entity). The sale of this product is expressly conditioned on the buyer not using the product or its components (1) in manufacturing; (2) to provide a service, information, or data to an unaffiliated third party for payment; (3) for therapeutic, diagnostic or prophylactic purposes; (4) to resell, sell, or otherwise transfer this product or its components to any third party, or for any other commercial purpose. Life Technologies Corporation will not assert a claim against the buyer of the infringement of the above patents based on the manufacture, use or sale of a commercial product developed in research by the buyer in which this product or its components was employed, provided that neither this product nor any of its components was used in the manufacture of such product. For information on purchasing a license to this product for purposes other than research, contact Life Technologies Corporation, Cell Analysis Business Unit, Business Development, 29851 Willow Creek Road, Eugene, OR 97402, Tel: (541) 465-8300. Fax: (541) 335-0354.

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