ANTIBODIES FOR COVID-19 RESEARCHKEY VIRAL AND HOST PROTEIN TARGETS

ANTIBODIES TO KEY VIRAL TARGETSSARS-CoV-2 is the causative agent of the global pandemic coronavirus disease 2019 (COVID-19). It shares ~80% of its RNA sequence with SARS-CoV, the identified pathogen of the 2002-2003 outbreak. Four major structural proteins have been identified: the nucleocapsid protein (N), spike protein (S), membrane protein (M) and envelope protein (E).

SARS-COV-2 NUCLEOCAPSID PROTEIN (N)

The SARS-COV Nucleocapsid (N) protein is highly immunogenic and expressed primarily in the cytoplasm. Probing for this protein is popularly used to confirm presence of the virus in tissue.

Notably, the SARS Nucleocapsid Protein Polyclonal Antibody (NB100-56576) from Novus Biologicals™ was recently validated by the US Centers for Disease Control and Prevention (CDC) for IHC Detection of SARS-CoV-2. Other recent publications of this antibody are listed below.

Pathology and pathogenesis of fatal COVID-19 cases associated with SARS-CoV-2 pandemic. Martines RB et al.; COVID-19 Pathology Working Group. Emerg Infect Dis. 2020 SARS Nucleocapsid Protein Antibody [NB100-56576] used for IHC.

Respiratory disease in rhesus macaques inoculated with SARS-CoV-2. Munster, V.J.et al.; National Institutes of Health, Nature 2020 [NB100-56576] used for IHC-P.

SARS-CoV-2 Infection Protects Against Rechallenge in Rhesus Macaques Chandrashekar A, et al.; Harvard Medical School, Science. 2020 [NB100-56576] used for IHC-P, IHC, Dual ISH-IHC.

In situ detection of SARS-CoV-2 in lungs and airways of patients with COVID-19. Schaefer, I., Padera, R.F. et al; Harvard Medical School, Mod Pathol. 2020 [NB100-56576] used for IHC.

Dr. Lynette Scholl, Harvard Medical School, recently published a study (Schaefer et al.) using this antibody and reported:

“We were able to quickly optimize and validate the antibody for use in clinical diagnostics on formalin fixed paraffin embedded tissues and have found that it is a robust tool for evaluation of SARS-CoV-2 in human autopsy tissue.”

SARS Nucleocapsid Protein Antibody [NB100-56576] - Immunostaining of SARS-CoV-2 in pulmonary tissues from fatal coronavirus disease cases.

A) P5 (Patient 5): immunostaining of tracheal epithelial cells. B) P5: immunostaining of ciliated cells. C) P8: immunostaining of desquamated type I pneumocyte in an alveolar lumen. D) P4: colocalization of SARS-CoV-2 viral antigen (red) with type II pneumocyte stained by surfactant (brown; arrow). E) P4: colocalization of SARS-CoV-2 viral antigen (red) with macrophages stained by CD163 (brown; arrows); virus immunostaining within type II pneumocytes is also seen (arrowheads). F) P4: immunostaining of hyaline membranes in a region of exudative DAD. G) P3: immunostaining within macrophage in hilar lymph node; anthracosis is also present. Emerg Infect Dis. 2020.

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ANTIBODIES TO KEY VIRAL TARGETSSARS SPIKE PROTEIN (S)

The spike protein (S) of SARS-CoV-2 is of major interest in COVID-19 research. It binds cellular membrane receptors, such as ACE-2, to initiate infection of the host cell. The S protein is composed of two functional subunits, S1 and S2, and requires priming by host proteases for entry. S1 contains a receptor binding domain (RBD) and is responsible for the initial attachment of the virus to the surface of host cells, and S2 is responsible for membrane fusion.

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LLAMABODYTM ANTIBODIES AND THE SARS-COV-2 SPIKE PROTEIN

Camelid antibodies are used in studying protein structure, high resolution imaging applications and drug delivery - as well as the detection and neutralization of viruses. Our SARS-CoV-1/2 Spike RBD Llamabody™ Antibody is a recombinant antibody with the VHH domain of the SARS VHH-72 clone attached to a human IgG scaffold. The small size of the VHH domain can reach more difficult epitopes, and this product is able to block both SARS-CoV-1 and SARS-CoV-2.

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SIMPLE WESTERN™ CERTIFIED ANTIBODIES FOR SARS-COV-2

We have antibodies for both SARS Nucleocapsid protein and SARS Spike protein that have been validated for ProteinSimple’s Simple WesternTM platforms, so are ready to slot into your workflow.

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SARS-CoV-2 Spike RBD Binding to ACE-2-transfected HEK293 Human Cell Line is Blocked by SARS-Cov-1/2 Spike RBD LlaMABodyTM In a functional flow cytometry test, His-tagged recombinant SARS-CoV-2 Spike RBD (50 ng/mL, 10534-CV) binds to HEK293 human embryonic kidney cell line transfected with human ACE-2 (black dotted line). Binding is completely blocked (orange histogram) by 25 µg/mL of Anti-SARS CoV-1/2 S1 RBD Llamabody (Catalog # LMAB10541). Mouse anti-Human DC-SIGN Monoclonal Antibody (MAB161) at 25 µg/mL was used as an irrelevant control (blue line). Cells were stained with Allophycocyanin-conjugated Mouse anti-His tag Monoclonal Antibody (IC050A).

(Left) Simple Western analysis of recombinant SARS-CoV-2 Nucleocapsid Protein (10474-CV) with SARS Nucleocapsid Protein Antibody [NB100-56683]. SARS Nucleocapsid protein was loaded at 20 ng/mL and detected using serial dilutions of the Rabbit Anti-SARS-CoV Nucleocapsid Protein Polyclonal Antibody (NB100-56683) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody.

(Right) Simple Western analysis of SARS-CoV-2 (1:50), MERS (1:100), OC43 (1:100), and 229E (1:100) lysates. A specific band was detected for SARS-CoV-2 Nucleocapsid Protein only in the SARS-CoV-2 lysate. Detection was based on the use of Rabbit Anti-SARS-CoV Nucleocapsid Protein Polyclonal Antibody [NB100-56683] followed by HRP-conjugated Anti-Goat IgG Secondary Antibody. Note: some reactivity observed with FL Std 230. SARS-CoV-2 lysate courtesy of University of Maryland. These experiments were conducted under reducing conditions and using the 12-230 kDa separation system.

ANTIBODIES TO KEY HUMAN TARGETSFrom research on SARS-host protein interactions, key human targets have been identified for their involvement in viral pathogenicity. These include cell surface receptors like ACE-2 and Neuropilin-1, as well as proteases such as TMPRSS2 and Furin.

ACE-2

Research on SARS-host protein interactions has shown that the viral spike (S) protein binds human ACE-2 receptor, resulting in fusion of viral and cell membranes for viral entry.

Our monoclonal Human ACE-2 Antibody (MAB933) was recently validated for IHC and Western blot by the Human Protein Atlas project. Our polyclonal Human ACE-2 Antibody (AF933) has been published in some of the latest research pertaining to SARS-CoV-2 viral entry.

The protein expression profile of ACE2 in human tissues Hikmet, F. et al., Human Protein Atlas Project, bioRxiv, 2020 Human ACE-2 Monoclonal Antibody (MAB933) used for IHC and WB

SARS-CoV-2 Cell Entry Depends on ACE2 and TMPRSS2 and Is Blocked by a Clinically Proven Protease Inhibitor. Hoffmann M. et al.; Leibniz Institute for Primate Research, Cell, 2020 Human ACE-2 Polyclonal Antibody [AF933] used for blocking.

SARS-CoV-2 productively infects human gut enterocytes Lamers M M. et al.; Hubrecht Institute, Science, 2020 Human ACE-2 Polyclonal Antibody [AF933] used for IHC.

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TMPRSS2 AND TMPRSS4

For the virus to enter the host cell, the S protein requires cleaving and activating by host cell proteases such as TMPRSS2 and TMPRSS4.

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FURIN

The enzyme furin has been protein of interest in SARS-CoV-2 research, since it was found that the viral S protein contains a furin-cleavage-site (FCS) in its sequence.

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NEUROPILIN-1

Neuropilin 1 (NRP1), also known as CD304, is a cell surface receptor that is known to bind furin-cleaved substrates. It is able to interact with the SARS-CoV-2 spike protein and enhance viral infection of the cell.

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Immunohistochemistry: ACE 2 in Hamster Lung. ACE 2 was detected in immersion fixed paraffin-embedded sections of hamster lung using Mouse Anti-Human ACE 2 Monoclonal Antibody (Catalog # MAB9332) at 10 µg/mL for 1 hour at room temperature followed by incubation with the Anti-Mouse IgG VisUCyte™ HRP Polymer Antibody (VC001). Before incubation with the primary antibody, tissue was subjected to heat-induced epitope retrieval using Antigen Retrieval Reagent-Basic (CTS013). Tissue was stained using DAB (brown) and counterstained with hematoxylin (blue). Specific staining was localized to respiratory bronchioles. Staining was performed using our protocol for IHC Staining with VisUCyte HRP Polymer Detection Reagents.

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ANTIBODIES TO KEY HUMAN TARGETS

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SARS-CoV-2 research is evolving at an extraordinary pace and new proteins of interest are being identified all the time. Below are just some of the other human targets that are being investigated for their role in SARS-CoV-2 replication and disease.

CATHEPSIN B/L Cathepsins B and L are other proteases that may be involved in priming of the SARS-CoV-2 S protein and facilitating infection of the cell.

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PIKFYVE Phosphatidylinositol 3-phosphate 5-kinase (PIKfyve) is an enzyme that synthesizes a key phosphoinositide (PI(3,5)P2) involved in regulating endosome dynamics. Inhibition of this enzyme may reduce SARS-CoV-2 infection.

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CD147 The transmembrane protein CD147 or EMPRINN has been proposed as an alternate receptor to ACE-2 for viral invasion.

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RDRP RNA-dependent RNA polymerase (RdRP) is an enzyme critical to the replication of RNA viruses such as SARS-CoV-2.

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IGG, IGA, IGM Antibodies specific to other immunoglobulins (Igs) are necessary to probe for their presence in serum, or to detect your primary antibody reagent.

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COVID-19 RESEARCH RESOURCES FROM NOVUS BIOLOGICALS

FLOW CYTOMETRY WORKFLOWS FOR COVID-19 RESEARCH

RECOMBINANT PROTEINS FOR CORONAVIRUS RESEARCH

SMALL MOLECULES FOR COVID-19 RESEARCH

FLOW CYTOMETRY IHC/ICC NEUTRALIZATION WESTERN BLOT SIMPLE WESTERN™ MATCHED PAIRS