Western blot analysis of extracts from mock-infected Vero-E6 cells (lane 1), SARS-CoV-2-infected Vero-E6 cells (lane 2), and isolated SARS-CoV-2 virions (lane 3), using SARS-CoV-2 Spike Protein (RBD) (E2T6M) Mouse mAb (upper) and GAPDH (D16H11) XP® Rabbit mAb #5174 (lower). The antibody detects full-length (uncleaved) SARS-CoV-2 spike protein, and the fragment corresponding to the S1 domain generated by endogenous protease cleavage. SARS-CoV-2 virions and SARS-CoV-2-infected Vero-E6 cells courtesy of Dr. Mohsan Saeed, National Emerging Infectious Diseases Laboratories (NEIDL), Boston University.
Western blot analysis of extracts from 293T cells, mock transfected (lane 1) or transiently transfected with expression constructs encoding Myc/DDK-tagged SARS-CoV-2 Spike (lane 2), Myc/DDK-tagged SARS-CoV Spike (lane 3), or Myc/DDK-tagged MERS-CoV Spike (lane 4), using SARS-CoV-2 Spike Protein (RBD) (E2T6M) Mouse mAb (upper), Myc-Tag (71D10) Rabbit mAb #2278 (middle), and GAPDH (D16H11) XP® Rabbit mAb #5174 (lower). The antibody detects full-length (uncleaved) SARS-CoV-2 spike protein, and the fragment corresponding to the S1 domain generated by endogenous protease cleavage. Proteolytic cleavage of SARS-CoV spike protein is not observed in these experimental conditions (transient transfection).
Western blot analysis of purified SARS-CoV-2 Spike RBD (318-541) Recombinant Protein (8xHis-Tag) #48801 (lane 1), SARS-CoV-2 Spike RBD (multimeric) (319-591) Recombinant Protein (8xHis-Tag) #17862 (lane 2), SARS-CoV-2 Spike (trimeric) (16-1208) Recombinant Protein (8xHis-Tag) #65444 (lane 3), and SARS-CoV-2 Spike S1-NTD (16-316) Recombinant Protein (8xHis-Tag) #88587 (lane 4), using SARS-CoV-2 Spike Protein (RBD) (E2T6M) Mouse mAb (upper) and His-Tag (D3I1O) XP® Rabbit mAb #12698 (lower). Due to the location of the epitope (surrounding Ser459 of SARS-CoV-2 spike protein), the antibody detects recombinant proteins corresponding to the full-length ectodomain and the receptor binding domain (RBD) of SARS CoV-2 Spike protein but does not detect recombinant proteins corresponding only to the SARS-CoV-2 S1 amino terminal domain (S1-NTD).
|MW (kDa)||110, 220|
|Source/Isotype||Mouse IgG1 kappa|
Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.
For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.
NOTE: Please refer to primary antibody product webpage for recommended antibody dilution.
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.
Load 20 µl onto SDS-PAGE gel (10 cm x 10 cm).
NOTE: Volumes are for 10 cm x 10 cm (100 cm2) of membrane; for different sized membranes, adjust volumes accordingly.
* Avoid repeated exposure to skin.
posted June 2005
revised June 2020
Protocol Id: 262
SARS-CoV-2 Spike Protein (RBD) (E2T6M) Mouse mAb recognizes endogenous levels of total SARS-CoV-2 spike protein. This antibody detects full-length protein, and also detects the S1 fragment generated by furin cleavage. It does not cross-react with spike proteins from SARS or MERS coronaviruses.
Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Ser459 of SARS-CoV-2 spike protein.
The cause of the COVID-19 pandemic is a novel and highly pathogenic coronavirus, termed SARS-CoV-2 (severe acute respiratory syndrome coronavirus-2). SARS-CoV-2 is a member of the Coronaviridae family of viruses (1). The genome of SARS-CoV-2 is similar to other coronaviruses, and is comprised of four key structural proteins: S, the spike protein, E, the envelope protein, M, the membrane protein, and N, the nucleocapsid protein (2). Coronavirus spike proteins are class I fusion proteins and harbor an ectodomain, a transmembrane domain, and an intracellular tail (3,4). The highly glycosylated ectodomain projects from the viral envelope surface and facilitates attachment and fusion with the host cell plasma membrane. The ectodomain can be further subdivided into host receptor-binding domain (RBD) (S1) and membrane-fusion (S2) subunits, which are produced upon proteolysis by host proteases at S1/S2 and S2’ sites. S1 and S2 subunits remain associated after cleavage and assemble into crown-like homotrimers (2,4). In humans, both SARS-CoV and SARS-CoV-2 spike proteins utilize the angiotensin-converting enzyme 2 (ACE2) protein as a receptor for cellular entry (5-7). Spike protein subunits represent a key antigenic feature of coronavirus virions, and therefore represent an important target of vaccines, novel therapeutic antibodies, and small-molecule inhibitors (8,9).
Explore pathways + proteins related to this product.
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