mers cov s rabbit polyclonal antibodies  (Sino Biological)


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    Name:
    MERS CoV Spike Protein Antibody Rabbit PAb
    Description:
    Produced in rabbits immunized with purified recombinant MERS CoV NCoV Novel coronavirus Spike Protein ECD aa 1 1297 Catalog 40069 V08B AFS88936 1 Met1 Trp1297 MERS CoV NCoV Novel coronavirus Spike Protein ECD aa 1 1297 specific IgG was purified by MERS CoV NCoV Novel coronavirus Spike Protein ECD aa 1 1297 affinity chromatography
    Catalog Number:
    40069-T30
    Price:
    None
    Category:
    Primary Antibody
    Reactivity:
    MERS CoV
    Applications:
    ELISA
    Immunogen:
    Recombinant MERS-CoV (NCoV / Novel coronavirus) Spike Protein (ECD, aa 1-1297) Protein (Catalog#40069-V08B)
    Product Aliases:
    Anti-coronavirus s1 Antibody, Anti-coronavirus s2 Antibody, Anti-coronavirus spike Antibody, Anti-cov spike Antibody, Anti-ncov RBD Antibody, Anti-ncov s1 Antibody, Anti-ncov s2 Antibody, Anti-ncov spike Antibody, Anti-RBD Antibody, Anti-S Antibody, Anti-s1 Antibody, Anti-Spike RBD Antibody
    Antibody Type:
    PAb
    Host:
    Rabbit
    Isotype:
    Rabbit IgG
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    Structured Review

    Sino Biological mers cov s rabbit polyclonal antibodies
    Two Selected G2-escape Mutations and Their Impact on G2 Binding and Neutralization. (A-D) Binding of G2 Fab to immobilized (A) <t>MERS-CoV</t> S1-NTD, (B) MERS-CoV S-2P, (C) MERS-CoV S-2P-S28F and (D) MERS-CoV S-2P-G198D measured by surface plasmon resonance (SPR). The same concentration series of G2 Fab was used in A-D. Best global fit of the data to a 1:1 binding model is shown as colored lines. (E) Neutralization activity of G2 IgG was measured against pseudotyped lentivirus bearing MERS-CoV S (WT) and two variants (S28F and G198D). Percent neutralization of WT (red), S28F (blue) and G198D (black) S pseudovirions at the different antibody concentrations is shown. Data points represent the mean of three technical replicates with standard errors. (F) Neutralization activity of G2 IgG was measured against authentic MERS-CoV (WT) and the G198D variant. Percent neutralization of WT (red) and G198D (black) MERS-CoV at the different antibody concentrations is shown. Data points for the wild-type virus represent the mean of two technical replicates.
    Produced in rabbits immunized with purified recombinant MERS CoV NCoV Novel coronavirus Spike Protein ECD aa 1 1297 Catalog 40069 V08B AFS88936 1 Met1 Trp1297 MERS CoV NCoV Novel coronavirus Spike Protein ECD aa 1 1297 specific IgG was purified by MERS CoV NCoV Novel coronavirus Spike Protein ECD aa 1 1297 affinity chromatography
    https://www.bioz.com/result/mers cov s rabbit polyclonal antibodies/product/Sino Biological
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    mers cov s rabbit polyclonal antibodies - by Bioz Stars, 2021-06
    94/100 stars

    Images

    1) Product Images from "Structural Definition of a Neutralization-sensitive Epitope on the MERS-CoV S1-NTD"

    Article Title: Structural Definition of a Neutralization-sensitive Epitope on the MERS-CoV S1-NTD

    Journal: Cell reports

    doi: 10.1016/j.celrep.2019.08.052

    Two Selected G2-escape Mutations and Their Impact on G2 Binding and Neutralization. (A-D) Binding of G2 Fab to immobilized (A) MERS-CoV S1-NTD, (B) MERS-CoV S-2P, (C) MERS-CoV S-2P-S28F and (D) MERS-CoV S-2P-G198D measured by surface plasmon resonance (SPR). The same concentration series of G2 Fab was used in A-D. Best global fit of the data to a 1:1 binding model is shown as colored lines. (E) Neutralization activity of G2 IgG was measured against pseudotyped lentivirus bearing MERS-CoV S (WT) and two variants (S28F and G198D). Percent neutralization of WT (red), S28F (blue) and G198D (black) S pseudovirions at the different antibody concentrations is shown. Data points represent the mean of three technical replicates with standard errors. (F) Neutralization activity of G2 IgG was measured against authentic MERS-CoV (WT) and the G198D variant. Percent neutralization of WT (red) and G198D (black) MERS-CoV at the different antibody concentrations is shown. Data points for the wild-type virus represent the mean of two technical replicates.
    Figure Legend Snippet: Two Selected G2-escape Mutations and Their Impact on G2 Binding and Neutralization. (A-D) Binding of G2 Fab to immobilized (A) MERS-CoV S1-NTD, (B) MERS-CoV S-2P, (C) MERS-CoV S-2P-S28F and (D) MERS-CoV S-2P-G198D measured by surface plasmon resonance (SPR). The same concentration series of G2 Fab was used in A-D. Best global fit of the data to a 1:1 binding model is shown as colored lines. (E) Neutralization activity of G2 IgG was measured against pseudotyped lentivirus bearing MERS-CoV S (WT) and two variants (S28F and G198D). Percent neutralization of WT (red), S28F (blue) and G198D (black) S pseudovirions at the different antibody concentrations is shown. Data points represent the mean of three technical replicates with standard errors. (F) Neutralization activity of G2 IgG was measured against authentic MERS-CoV (WT) and the G198D variant. Percent neutralization of WT (red) and G198D (black) MERS-CoV at the different antibody concentrations is shown. Data points for the wild-type virus represent the mean of two technical replicates.

    Techniques Used: Binding Assay, Neutralization, SPR Assay, Concentration Assay, Activity Assay, Variant Assay

    G2 IgG Prevents the Binding of MERS-CoV S Protein to DDP4-Expressing Cells. Normalized binding efficiency of GFP-tagged MERS-CoV S-2P proteins to DPP4-expressing FreeStyle 293F cells in the presence or absence of IgGs was calculated from median fluorescence intensity (MFI) values. FreeStyle 293-F cells were transfected with a plasmid encoding full-length DPP4 60 h before the experiment. Non-transfected cells (NT) incubated with MERS-CoV S-2P, as well as transfected cells incubated with PBS, were used as negative controls. AM14 is an irrelevant RSV F-specific neutralizing antibody used as another negative control. Bar graph shows the mean and error bars indicate the standard deviation (n = 3 biologically independent experiments with two technical replicates).
    Figure Legend Snippet: G2 IgG Prevents the Binding of MERS-CoV S Protein to DDP4-Expressing Cells. Normalized binding efficiency of GFP-tagged MERS-CoV S-2P proteins to DPP4-expressing FreeStyle 293F cells in the presence or absence of IgGs was calculated from median fluorescence intensity (MFI) values. FreeStyle 293-F cells were transfected with a plasmid encoding full-length DPP4 60 h before the experiment. Non-transfected cells (NT) incubated with MERS-CoV S-2P, as well as transfected cells incubated with PBS, were used as negative controls. AM14 is an irrelevant RSV F-specific neutralizing antibody used as another negative control. Bar graph shows the mean and error bars indicate the standard deviation (n = 3 biologically independent experiments with two technical replicates).

    Techniques Used: Binding Assay, Expressing, Fluorescence, Transfection, Plasmid Preparation, Incubation, Negative Control, Standard Deviation

    G2-mediated Inhibition of DPP4 binding to MERS-CoV S Depends on Oligomeric State of the Spike (A–C) Normalized binding efficiency of DPP4, in the presence or absence of Fab, to cells transfected with plasmids encoding (A) membrane-anchored S1 (S1-TM), (B) full-length S-WT (S-WT-FL) and (C) full-length S-2P (S-2P-FL). Cells incubated with PBS were used as negative controls. Bar graph shows the mean and error bars indicate the standard deviation (n = 3 biologically independent experiments with two technical replicates). (D) Surface plasmon resonance competition assay. Response curves for MERS-CoV S-2P, alone or in the presence of a 5-fold molar excess of indicated Fabs or IgGs, passed over immobilized DPP4 ectodomain. The curves for S-2P or S-2P supplemented with Fab or IgG are shown with a solid line, whereas control curves for samples without S-2P are shown with a dotted line.
    Figure Legend Snippet: G2-mediated Inhibition of DPP4 binding to MERS-CoV S Depends on Oligomeric State of the Spike (A–C) Normalized binding efficiency of DPP4, in the presence or absence of Fab, to cells transfected with plasmids encoding (A) membrane-anchored S1 (S1-TM), (B) full-length S-WT (S-WT-FL) and (C) full-length S-2P (S-2P-FL). Cells incubated with PBS were used as negative controls. Bar graph shows the mean and error bars indicate the standard deviation (n = 3 biologically independent experiments with two technical replicates). (D) Surface plasmon resonance competition assay. Response curves for MERS-CoV S-2P, alone or in the presence of a 5-fold molar excess of indicated Fabs or IgGs, passed over immobilized DPP4 ectodomain. The curves for S-2P or S-2P supplemented with Fab or IgG are shown with a solid line, whereas control curves for samples without S-2P are shown with a dotted line.

    Techniques Used: Inhibition, Binding Assay, Transfection, Incubation, Standard Deviation, SPR Assay, Competitive Binding Assay

    2) Product Images from "A high-throughput inhibition assay to study MERS-CoV antibody interactions using image cytometry"

    Article Title: A high-throughput inhibition assay to study MERS-CoV antibody interactions using image cytometry

    Journal: Journal of virological methods

    doi: 10.1016/j.jviromet.2018.11.009

    Ability of mAbs to inhibit MERS-CoV S protein binding to DPP4-expressing cells. Inhibition of MERS-CoV S binding to DPP4 by D12 mAb was measured by image cytometry (A) and fluorescence intensity was analyzed by FlowJo histograms (B). As a control, a non-relevant isotype monoclonal Ab was used. Dose response curves of Log mAb concentration versus percent inhibition were generated for D12 (C), G2 (D), and G4 (E) mAbs. (C-E) Each dot represents duplicates. Error bars represent SEM.
    Figure Legend Snippet: Ability of mAbs to inhibit MERS-CoV S protein binding to DPP4-expressing cells. Inhibition of MERS-CoV S binding to DPP4 by D12 mAb was measured by image cytometry (A) and fluorescence intensity was analyzed by FlowJo histograms (B). As a control, a non-relevant isotype monoclonal Ab was used. Dose response curves of Log mAb concentration versus percent inhibition were generated for D12 (C), G2 (D), and G4 (E) mAbs. (C-E) Each dot represents duplicates. Error bars represent SEM.

    Techniques Used: Protein Binding, Expressing, Inhibition, Binding Assay, Cytometry, Fluorescence, Concentration Assay, Generated

    MERS-CoV S1 and S proteins binding to DPP4-expressing cells. Whole well fluorescence images of MERS-CoV S1 (A) and MERS-CoV S (B) proteins binding DPP4-expressing BHK21 cells. Both S1 monomer and S trimer proteins were stained with anti-MERSCoV S polyclonal AF488, demonstrating binding to DPP4-expressing cells in green. Analysis of the image-based fluorescence intensity data by FlowJo for MERS-CoV S1 monomer (C) and S trimer (D) shows the peak shift results from binding.
    Figure Legend Snippet: MERS-CoV S1 and S proteins binding to DPP4-expressing cells. Whole well fluorescence images of MERS-CoV S1 (A) and MERS-CoV S (B) proteins binding DPP4-expressing BHK21 cells. Both S1 monomer and S trimer proteins were stained with anti-MERSCoV S polyclonal AF488, demonstrating binding to DPP4-expressing cells in green. Analysis of the image-based fluorescence intensity data by FlowJo for MERS-CoV S1 monomer (C) and S trimer (D) shows the peak shift results from binding.

    Techniques Used: Binding Assay, Expressing, Fluorescence, Staining

    3) Product Images from "Structural Definition of a Neutralization-Sensitive Epitope on the MERS-CoV S1-NTD"

    Article Title: Structural Definition of a Neutralization-Sensitive Epitope on the MERS-CoV S1-NTD

    Journal: Cell Reports

    doi: 10.1016/j.celrep.2019.08.052

    G2 IgG Prevents the Binding of MERS-CoV S Protein to DDP4-Expressing Cells Normalized binding efficiency of GFP-tagged MERS-CoV S-2P proteins to DPP4-expressing FreeStyle 293F cells in the presence or absence of IgGs was calculated from median fluorescence intensity (MFI) values. FreeStyle 293-F cells were transfected with a plasmid encoding full-length DPP4 60 h before the experiment. Non-transfected cells (NTs) incubated with MERS-CoV S-2P, as well as transfected cells incubated with PBS, were used as negative controls. AM14 is an irrelevant RSV F-specific neutralizing antibody used as another negative control. Bar graph shows the mean and error bars indicate the standard deviation (n = 3 biologically independent experiments with two technical replicates).
    Figure Legend Snippet: G2 IgG Prevents the Binding of MERS-CoV S Protein to DDP4-Expressing Cells Normalized binding efficiency of GFP-tagged MERS-CoV S-2P proteins to DPP4-expressing FreeStyle 293F cells in the presence or absence of IgGs was calculated from median fluorescence intensity (MFI) values. FreeStyle 293-F cells were transfected with a plasmid encoding full-length DPP4 60 h before the experiment. Non-transfected cells (NTs) incubated with MERS-CoV S-2P, as well as transfected cells incubated with PBS, were used as negative controls. AM14 is an irrelevant RSV F-specific neutralizing antibody used as another negative control. Bar graph shows the mean and error bars indicate the standard deviation (n = 3 biologically independent experiments with two technical replicates).

    Techniques Used: Binding Assay, Expressing, Fluorescence, Transfection, Plasmid Preparation, Incubation, Negative Control, Standard Deviation

    Two Selected G2-Escape Mutations and Their Impact on G2 Binding and Neutralization (A–D) Binding of G2 Fab to immobilized (A) MERS-CoV S1-NTD, (B) MERS-CoV S-2P, (C) MERS-CoV S-2P-S28F, and (D) MERS-CoV S-2P-G198D measured by surface plasmon resonance (SPR). The same concentration series of G2 Fab was used in (A)–(D). Best global fit of the data to a 1:1 binding model is shown as colored lines. (E) Neutralization activity of G2 IgG was measured against pseudotyped lentivirus bearing MERS-CoV S (WT) and two variants (S28F and G198D). Percent neutralization of WT (red), S28F (blue), and G198D (black) S pseudovirions at the different antibody concentrations is shown. Data points represent the mean of three technical replicates with standard errors. (F) Neutralization activity of G2 IgG was measured against authentic MERS-CoV (WT) and the G198D variant. Percent neutralization of WT (red) and G198D (black) MERS-CoV at the different antibody concentrations is shown. Data points for the WT virus represent the mean of two technical replicates.
    Figure Legend Snippet: Two Selected G2-Escape Mutations and Their Impact on G2 Binding and Neutralization (A–D) Binding of G2 Fab to immobilized (A) MERS-CoV S1-NTD, (B) MERS-CoV S-2P, (C) MERS-CoV S-2P-S28F, and (D) MERS-CoV S-2P-G198D measured by surface plasmon resonance (SPR). The same concentration series of G2 Fab was used in (A)–(D). Best global fit of the data to a 1:1 binding model is shown as colored lines. (E) Neutralization activity of G2 IgG was measured against pseudotyped lentivirus bearing MERS-CoV S (WT) and two variants (S28F and G198D). Percent neutralization of WT (red), S28F (blue), and G198D (black) S pseudovirions at the different antibody concentrations is shown. Data points represent the mean of three technical replicates with standard errors. (F) Neutralization activity of G2 IgG was measured against authentic MERS-CoV (WT) and the G198D variant. Percent neutralization of WT (red) and G198D (black) MERS-CoV at the different antibody concentrations is shown. Data points for the WT virus represent the mean of two technical replicates.

    Techniques Used: Binding Assay, Neutralization, SPR Assay, Concentration Assay, Activity Assay, Variant Assay

    G2-Mediated Inhibition of DPP4 Binding to MERS-CoV S Depends on the Oligomeric State of the Spike (A–C) Normalized binding efficiency of DPP4, in the presence or absence of Fab, to cells transfected with plasmids encoding (A) membrane-anchored S1 (S1-TM), (B) full-length S-WT (S-WT-FL), and (C) full-length S-2P (S-2P-FL). Cells incubated with PBS were used as negative controls. Bar graph shows the mean and error bars indicate the standard deviation (n = 3 biologically independent experiments with two technical replicates). (D) Surface plasmon resonance competition assay. Response curves for MERS-CoV S-2P, alone or in the presence of a 5-fold molar excess of indicated Fabs or IgGs, passed over an immobilized DPP4 ectodomain. The curves for S-2P or S-2P supplemented with Fab or IgG are shown with a solid line, whereas control curves for samples without S-2P are shown with a dotted line.
    Figure Legend Snippet: G2-Mediated Inhibition of DPP4 Binding to MERS-CoV S Depends on the Oligomeric State of the Spike (A–C) Normalized binding efficiency of DPP4, in the presence or absence of Fab, to cells transfected with plasmids encoding (A) membrane-anchored S1 (S1-TM), (B) full-length S-WT (S-WT-FL), and (C) full-length S-2P (S-2P-FL). Cells incubated with PBS were used as negative controls. Bar graph shows the mean and error bars indicate the standard deviation (n = 3 biologically independent experiments with two technical replicates). (D) Surface plasmon resonance competition assay. Response curves for MERS-CoV S-2P, alone or in the presence of a 5-fold molar excess of indicated Fabs or IgGs, passed over an immobilized DPP4 ectodomain. The curves for S-2P or S-2P supplemented with Fab or IgG are shown with a solid line, whereas control curves for samples without S-2P are shown with a dotted line.

    Techniques Used: Inhibition, Binding Assay, Transfection, Incubation, Standard Deviation, SPR Assay, Competitive Binding Assay

    Related Articles

    Incubation:

    Article Title: Characterization of a human monoclonal antibody generated from a B-cell specific for a prefusion-stabilized spike protein of Middle East respiratory syndrome coronavirus
    Article Snippet: Then, human anti-MERS-CoV antibodies were incubated with the Vero cells infected with MERS-CoV. .. As positive controls, a rabbit polyclonal anti-MERS-CoV spike protein antibody (Sino biological Inc., Beijing, China) for MERS-CoV, a mouse anti-229E coronavirus nucleoprotein OC-43 antibody (MERCK, Darmstadt, Germany) for HCoV-229E, a mouse anti-coronavirus antibody, hCoV OC-43 (LifeSpan BioScience, Seattle, WA, USA) for HCoV-OC43, a rabbit polyclonal anti-hCoV-HKU1 spike protein antibody (Sino Biological Inc., Beijing, China) for hCoV-NL63, a human serum of a Korean SARS-CoV-2 convalescent person were also incubated. .. The secondary antibodies, Fluorescein isothiocyanate (FITC) -conjugated goat anti-human-IgG (Abcam, Cambridge, U.K.), FITC-conjugated goat anti-rabbit-IgG (Abcam, Cambridge, U.K.), and FITC-conjugated goat anti-mouse-IgG (Jackson Immunoresearch, West Grove, PA, USA) were added to the cells and incubated.

    Article Title: A Novel Bacterium-Like Particle Vaccine Displaying the MERS-CoV Receptor-Binding Domain Induces Specific Mucosal and Systemic Immune Responses in Mice
    Article Snippet: Briefly, Sf9 cells cultured in 96-well plates at 2 × 106 cells/mL were infected with the recombinant baculovirus. .. After 48 h of infection, the cultured plates were fixed with 80% cold acetone overnight at -20°C, washed three times with PBS-0.05% Tween 20 (PBST), and then incubated with a rabbit anti-MERS-S polyclonal antibody (1:500, Sino Biological Inc, Beijing, China) containing 1% bovine serum albumin (BSA, Sigma-Aldrich, USA) at 37°C for 1 h. After three washes with PBST, an FITC-labeled goat against rabbit IgG antibody (1:300, BioWorld, Inc, St. Louis, MN, USA) was added with Evans blue (Sigma-Aldrich, St. Louis, MN, USA) for 1 h at 37 °C. ..

    Article Title: A high-throughput inhibition assay to study MERS-CoV antibody interactions using image cytometry
    Article Snippet: After incubation, cells were washed with PBS, fixed with 80% cold acetone for 20 min at RT, and washed again with PBS. .. Subsequently, 100 μL of MERS-CoV S rabbit polyclonal antibodies (Sino Biological, Beijing, China) was pipetted into each well, incubated for 60 min at RT, and washed twice. .. Next, secondary goat anti-rabbit IgG H & L labeled with Alexa Fluor® 488 (AF488) and DPP4 PE-conjugated mouse antibody (Sino Biological) were added, incubated for 60 min at RT, and washed twice.

    Infection:

    Article Title: A Novel Bacterium-Like Particle Vaccine Displaying the MERS-CoV Receptor-Binding Domain Induces Specific Mucosal and Systemic Immune Responses in Mice
    Article Snippet: Briefly, Sf9 cells cultured in 96-well plates at 2 × 106 cells/mL were infected with the recombinant baculovirus. .. After 48 h of infection, the cultured plates were fixed with 80% cold acetone overnight at -20°C, washed three times with PBS-0.05% Tween 20 (PBST), and then incubated with a rabbit anti-MERS-S polyclonal antibody (1:500, Sino Biological Inc, Beijing, China) containing 1% bovine serum albumin (BSA, Sigma-Aldrich, USA) at 37°C for 1 h. After three washes with PBST, an FITC-labeled goat against rabbit IgG antibody (1:300, BioWorld, Inc, St. Louis, MN, USA) was added with Evans blue (Sigma-Aldrich, St. Louis, MN, USA) for 1 h at 37 °C. ..

    Cell Culture:

    Article Title: A Novel Bacterium-Like Particle Vaccine Displaying the MERS-CoV Receptor-Binding Domain Induces Specific Mucosal and Systemic Immune Responses in Mice
    Article Snippet: Briefly, Sf9 cells cultured in 96-well plates at 2 × 106 cells/mL were infected with the recombinant baculovirus. .. After 48 h of infection, the cultured plates were fixed with 80% cold acetone overnight at -20°C, washed three times with PBS-0.05% Tween 20 (PBST), and then incubated with a rabbit anti-MERS-S polyclonal antibody (1:500, Sino Biological Inc, Beijing, China) containing 1% bovine serum albumin (BSA, Sigma-Aldrich, USA) at 37°C for 1 h. After three washes with PBST, an FITC-labeled goat against rabbit IgG antibody (1:300, BioWorld, Inc, St. Louis, MN, USA) was added with Evans blue (Sigma-Aldrich, St. Louis, MN, USA) for 1 h at 37 °C. ..

    Immunohistochemistry:

    Article Title: Intratracheal exposure of common marmosets to MERS-CoV Jordan-n3/2012 or MERS-CoV EMC/2012 isolates does not result in lethal disease
    Article Snippet: .. To detect MERS-CoV antigen, IHC was performed using a rabbit polyclonal antiserum against MERS-CoV (Sino Biological PR China) (1:1000). .. Tissues from an uninfected control animal were used to validate all IHC procedures.

    Recombinant:

    Article Title: Development of a SERS-based lateral flow immunoassay for rapid and ultra-sensitive detection of anti-SARS-CoV-2 IgM/IgG in clinical samples
    Article Snippet: 2.1 Materials and chemicals Tetraethyl orthosilicate (TEOS), 2-(N-morpholino) ethanesulfonic (MES), polyethyleneimine (PEI, 25 K), polyvinylpyrrolidone (PVP, 40 K), Tween 20, N-hydroxysuccinimide (NHS), N-(3-dimethylaminopropyl)-N′-ethylcarbodiimide hydrochloride (EDC), 5,5-dithiobis-(2-nitrobenzoic acid) (DTNB), sodium borohydride (NaBH4 ), trisodium citrate (Na3 C6 H5 O7 , TSC), goat anti-human IgM, goat anti-human IgG, goat anti-rabbit IgG, bovine serum albumin (BSA) were obtained from Sigma-Aldrich (USA). .. SARS-CoV-2 S protein (recombinant) and S protein antibody (Ab) were obtained from Sino Biological Inc. (Beijing, China). .. Chloroauric acid tetrahydrate, sodium borohydride, formaldehyde (37%, w/w), ammonia solution (28%, w/w), chloroauric acid tetrahydrate (HAuCl4 ·4H2 O), and silver nitrate (AgNO3) were obtained from Sinopharm Chemical Reagent Co., Ltd. (Shanghai, China).

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  • 94
    Sino Biological mers cov s rabbit polyclonal antibodies
    Two Selected G2-escape Mutations and Their Impact on G2 Binding and Neutralization. (A-D) Binding of G2 Fab to immobilized (A) <t>MERS-CoV</t> S1-NTD, (B) MERS-CoV S-2P, (C) MERS-CoV S-2P-S28F and (D) MERS-CoV S-2P-G198D measured by surface plasmon resonance (SPR). The same concentration series of G2 Fab was used in A-D. Best global fit of the data to a 1:1 binding model is shown as colored lines. (E) Neutralization activity of G2 IgG was measured against pseudotyped lentivirus bearing MERS-CoV S (WT) and two variants (S28F and G198D). Percent neutralization of WT (red), S28F (blue) and G198D (black) S pseudovirions at the different antibody concentrations is shown. Data points represent the mean of three technical replicates with standard errors. (F) Neutralization activity of G2 IgG was measured against authentic MERS-CoV (WT) and the G198D variant. Percent neutralization of WT (red) and G198D (black) MERS-CoV at the different antibody concentrations is shown. Data points for the wild-type virus represent the mean of two technical replicates.
    Mers Cov S Rabbit Polyclonal Antibodies, supplied by Sino Biological, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/mers cov s rabbit polyclonal antibodies/product/Sino Biological
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    mers cov s rabbit polyclonal antibodies - by Bioz Stars, 2021-06
    94/100 stars
      Buy from Supplier

    95
    Sino Biological mers cov nucleocapsid protein antibody rabbit pab
    IHC detection of virus antigen expression in mouse tissue after challenge with <t>MERS-CoV.</t> Lung (A–C) and trachea (D–F) sections were assessed using rabbit polyclonal antibody to MERS-CoV nucleoprotein (NP) 3 days after the MERS-CoV challenge. The dark purple spot marked the inflammatory cell infiltration, and the brown particle marked the antigen of MERS-CoV. The MERS-CoV was located mainly in the trachea. Additionally, the lung tissue showed MERS-CoV expression in all immunized groups.
    Mers Cov Nucleocapsid Protein Antibody Rabbit Pab, supplied by Sino Biological, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/mers cov nucleocapsid protein antibody rabbit pab/product/Sino Biological
    Average 95 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    mers cov nucleocapsid protein antibody rabbit pab - by Bioz Stars, 2021-06
    95/100 stars
      Buy from Supplier

    94
    Sino Biological mers cov spike antibody rabbit pab
    Pseudo-particle assays of <t>MERS-CoV</t> S WT and mutants. Huh-7 cells were infected with MLV-based pseudo-particles (PP) carrying MERS-CoV S WT or one of the respective S mutants. After 72 h, infected cells were lysed and assessed for luciferase activity. (A) PP infectivity of Huh 7 cells. (B) Infectivity of PP carrying the D922A S protein. Δenv and VSV-G served as representative controls for all PP assays (C) Impact of intracellular Ca 2+ on MERS-CoV fusion. Cells were pre-treated with growth medium containing either 50 µM calcium chelator BAPTA-AM ordimethyl sulfoxide (DMSO) for 1 h. Cells were then infected with their respective PP in the presence of BAPTA-AM or DMSO for 2 h and grown for 72 h before assessing for luciferase activity. (D) Impact of extracellular Ca 2+ on MERS-CoV fusion. Cells were pre-treated with growth medium either with or without 1.8 mM Ca 2+ for 1 h. Infection protocol is as described above except PP were treated with 1.5 mM EGTA for calcium chelation. Infectivity was normalized such that WT PP infectivity is 1. Error bars represent standard deviation (n = 3). Statistical analysis was performed using an unpaired student’s t-test, as indicated. * = p > 0.5, ** = p > 0.05, *** = p > 0.005.
    Mers Cov Spike Antibody Rabbit Pab, supplied by Sino Biological, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/mers cov spike antibody rabbit pab/product/Sino Biological
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    mers cov spike antibody rabbit pab - by Bioz Stars, 2021-06
    94/100 stars
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    Image Search Results


    Two Selected G2-escape Mutations and Their Impact on G2 Binding and Neutralization. (A-D) Binding of G2 Fab to immobilized (A) MERS-CoV S1-NTD, (B) MERS-CoV S-2P, (C) MERS-CoV S-2P-S28F and (D) MERS-CoV S-2P-G198D measured by surface plasmon resonance (SPR). The same concentration series of G2 Fab was used in A-D. Best global fit of the data to a 1:1 binding model is shown as colored lines. (E) Neutralization activity of G2 IgG was measured against pseudotyped lentivirus bearing MERS-CoV S (WT) and two variants (S28F and G198D). Percent neutralization of WT (red), S28F (blue) and G198D (black) S pseudovirions at the different antibody concentrations is shown. Data points represent the mean of three technical replicates with standard errors. (F) Neutralization activity of G2 IgG was measured against authentic MERS-CoV (WT) and the G198D variant. Percent neutralization of WT (red) and G198D (black) MERS-CoV at the different antibody concentrations is shown. Data points for the wild-type virus represent the mean of two technical replicates.

    Journal: Cell reports

    Article Title: Structural Definition of a Neutralization-sensitive Epitope on the MERS-CoV S1-NTD

    doi: 10.1016/j.celrep.2019.08.052

    Figure Lengend Snippet: Two Selected G2-escape Mutations and Their Impact on G2 Binding and Neutralization. (A-D) Binding of G2 Fab to immobilized (A) MERS-CoV S1-NTD, (B) MERS-CoV S-2P, (C) MERS-CoV S-2P-S28F and (D) MERS-CoV S-2P-G198D measured by surface plasmon resonance (SPR). The same concentration series of G2 Fab was used in A-D. Best global fit of the data to a 1:1 binding model is shown as colored lines. (E) Neutralization activity of G2 IgG was measured against pseudotyped lentivirus bearing MERS-CoV S (WT) and two variants (S28F and G198D). Percent neutralization of WT (red), S28F (blue) and G198D (black) S pseudovirions at the different antibody concentrations is shown. Data points represent the mean of three technical replicates with standard errors. (F) Neutralization activity of G2 IgG was measured against authentic MERS-CoV (WT) and the G198D variant. Percent neutralization of WT (red) and G198D (black) MERS-CoV at the different antibody concentrations is shown. Data points for the wild-type virus represent the mean of two technical replicates.

    Article Snippet: Subsequently, cells were stained with MERS-CoV S rabbit polyclonal antibodies (Sino Biological, Beijing, China) and then secondary goat anti-rabbit IgG H & L labeled with Alexa Fluor® 488 (AF488) was added.

    Techniques: Binding Assay, Neutralization, SPR Assay, Concentration Assay, Activity Assay, Variant Assay

    G2 IgG Prevents the Binding of MERS-CoV S Protein to DDP4-Expressing Cells. Normalized binding efficiency of GFP-tagged MERS-CoV S-2P proteins to DPP4-expressing FreeStyle 293F cells in the presence or absence of IgGs was calculated from median fluorescence intensity (MFI) values. FreeStyle 293-F cells were transfected with a plasmid encoding full-length DPP4 60 h before the experiment. Non-transfected cells (NT) incubated with MERS-CoV S-2P, as well as transfected cells incubated with PBS, were used as negative controls. AM14 is an irrelevant RSV F-specific neutralizing antibody used as another negative control. Bar graph shows the mean and error bars indicate the standard deviation (n = 3 biologically independent experiments with two technical replicates).

    Journal: Cell reports

    Article Title: Structural Definition of a Neutralization-sensitive Epitope on the MERS-CoV S1-NTD

    doi: 10.1016/j.celrep.2019.08.052

    Figure Lengend Snippet: G2 IgG Prevents the Binding of MERS-CoV S Protein to DDP4-Expressing Cells. Normalized binding efficiency of GFP-tagged MERS-CoV S-2P proteins to DPP4-expressing FreeStyle 293F cells in the presence or absence of IgGs was calculated from median fluorescence intensity (MFI) values. FreeStyle 293-F cells were transfected with a plasmid encoding full-length DPP4 60 h before the experiment. Non-transfected cells (NT) incubated with MERS-CoV S-2P, as well as transfected cells incubated with PBS, were used as negative controls. AM14 is an irrelevant RSV F-specific neutralizing antibody used as another negative control. Bar graph shows the mean and error bars indicate the standard deviation (n = 3 biologically independent experiments with two technical replicates).

    Article Snippet: Subsequently, cells were stained with MERS-CoV S rabbit polyclonal antibodies (Sino Biological, Beijing, China) and then secondary goat anti-rabbit IgG H & L labeled with Alexa Fluor® 488 (AF488) was added.

    Techniques: Binding Assay, Expressing, Fluorescence, Transfection, Plasmid Preparation, Incubation, Negative Control, Standard Deviation

    G2-mediated Inhibition of DPP4 binding to MERS-CoV S Depends on Oligomeric State of the Spike (A–C) Normalized binding efficiency of DPP4, in the presence or absence of Fab, to cells transfected with plasmids encoding (A) membrane-anchored S1 (S1-TM), (B) full-length S-WT (S-WT-FL) and (C) full-length S-2P (S-2P-FL). Cells incubated with PBS were used as negative controls. Bar graph shows the mean and error bars indicate the standard deviation (n = 3 biologically independent experiments with two technical replicates). (D) Surface plasmon resonance competition assay. Response curves for MERS-CoV S-2P, alone or in the presence of a 5-fold molar excess of indicated Fabs or IgGs, passed over immobilized DPP4 ectodomain. The curves for S-2P or S-2P supplemented with Fab or IgG are shown with a solid line, whereas control curves for samples without S-2P are shown with a dotted line.

    Journal: Cell reports

    Article Title: Structural Definition of a Neutralization-sensitive Epitope on the MERS-CoV S1-NTD

    doi: 10.1016/j.celrep.2019.08.052

    Figure Lengend Snippet: G2-mediated Inhibition of DPP4 binding to MERS-CoV S Depends on Oligomeric State of the Spike (A–C) Normalized binding efficiency of DPP4, in the presence or absence of Fab, to cells transfected with plasmids encoding (A) membrane-anchored S1 (S1-TM), (B) full-length S-WT (S-WT-FL) and (C) full-length S-2P (S-2P-FL). Cells incubated with PBS were used as negative controls. Bar graph shows the mean and error bars indicate the standard deviation (n = 3 biologically independent experiments with two technical replicates). (D) Surface plasmon resonance competition assay. Response curves for MERS-CoV S-2P, alone or in the presence of a 5-fold molar excess of indicated Fabs or IgGs, passed over immobilized DPP4 ectodomain. The curves for S-2P or S-2P supplemented with Fab or IgG are shown with a solid line, whereas control curves for samples without S-2P are shown with a dotted line.

    Article Snippet: Subsequently, cells were stained with MERS-CoV S rabbit polyclonal antibodies (Sino Biological, Beijing, China) and then secondary goat anti-rabbit IgG H & L labeled with Alexa Fluor® 488 (AF488) was added.

    Techniques: Inhibition, Binding Assay, Transfection, Incubation, Standard Deviation, SPR Assay, Competitive Binding Assay

    Sequencing of MERS-CoV Spike protein and data analysis

    Journal: Virology

    Article Title: Intratracheal exposure of common marmosets to MERS-CoV Jordan-n3/2012 or MERS-CoV EMC/2012 isolates does not result in lethal disease

    doi: 10.1016/j.virol.2015.07.013

    Figure Lengend Snippet: Sequencing of MERS-CoV Spike protein and data analysis

    Article Snippet: To detect MERS-CoV antigen, IHC was performed using a rabbit polyclonal antiserum against MERS-CoV (Sino Biological PR China) (1:1000).

    Techniques: Sequencing

    Gross and Histopathology. (A) Gross lung pathology demonstrating mostly normal lung with multifocal to coalescing moderate interstitial pneumonia in the left caudal lung lobe. (B) Low magnification of lung field from MERS-CoV EMC inoculated subject demonstrating

    Journal: Virology

    Article Title: Intratracheal exposure of common marmosets to MERS-CoV Jordan-n3/2012 or MERS-CoV EMC/2012 isolates does not result in lethal disease

    doi: 10.1016/j.virol.2015.07.013

    Figure Lengend Snippet: Gross and Histopathology. (A) Gross lung pathology demonstrating mostly normal lung with multifocal to coalescing moderate interstitial pneumonia in the left caudal lung lobe. (B) Low magnification of lung field from MERS-CoV EMC inoculated subject demonstrating

    Article Snippet: To detect MERS-CoV antigen, IHC was performed using a rabbit polyclonal antiserum against MERS-CoV (Sino Biological PR China) (1:1000).

    Techniques: Histopathology

    Ex-vivo analysis of primary cells. Cells were isolated from lung, kidney, and bronchoalveolar lavage (BAL), and one-step growth kinetics were performed as described in Materials and Methods. Lung, Kidney and BAL demonstrate that MERS-CoV is able to replicate

    Journal: Virology

    Article Title: Intratracheal exposure of common marmosets to MERS-CoV Jordan-n3/2012 or MERS-CoV EMC/2012 isolates does not result in lethal disease

    doi: 10.1016/j.virol.2015.07.013

    Figure Lengend Snippet: Ex-vivo analysis of primary cells. Cells were isolated from lung, kidney, and bronchoalveolar lavage (BAL), and one-step growth kinetics were performed as described in Materials and Methods. Lung, Kidney and BAL demonstrate that MERS-CoV is able to replicate

    Article Snippet: To detect MERS-CoV antigen, IHC was performed using a rabbit polyclonal antiserum against MERS-CoV (Sino Biological PR China) (1:1000).

    Techniques: Ex Vivo, Isolation

    Fluorescence Reduction Neutralizing Assay. Sera from all subjects was evaluated for the presence of neutralizing antibody to MERS-CoV as described in Materials and Methods. Neutralizing antibody could be detected above the background of the Mock infected

    Journal: Virology

    Article Title: Intratracheal exposure of common marmosets to MERS-CoV Jordan-n3/2012 or MERS-CoV EMC/2012 isolates does not result in lethal disease

    doi: 10.1016/j.virol.2015.07.013

    Figure Lengend Snippet: Fluorescence Reduction Neutralizing Assay. Sera from all subjects was evaluated for the presence of neutralizing antibody to MERS-CoV as described in Materials and Methods. Neutralizing antibody could be detected above the background of the Mock infected

    Article Snippet: To detect MERS-CoV antigen, IHC was performed using a rabbit polyclonal antiserum against MERS-CoV (Sino Biological PR China) (1:1000).

    Techniques: Fluorescence, Neutralizing Assay, Infection

    IHC detection of virus antigen expression in mouse tissue after challenge with MERS-CoV. Lung (A–C) and trachea (D–F) sections were assessed using rabbit polyclonal antibody to MERS-CoV nucleoprotein (NP) 3 days after the MERS-CoV challenge. The dark purple spot marked the inflammatory cell infiltration, and the brown particle marked the antigen of MERS-CoV. The MERS-CoV was located mainly in the trachea. Additionally, the lung tissue showed MERS-CoV expression in all immunized groups.

    Journal: Vaccine

    Article Title: The recombinant N-terminal domain of spike proteins is a potential vaccine against Middle East respiratory syndrome coronavirus (MERS-CoV) infection

    doi: 10.1016/j.vaccine.2016.11.064

    Figure Lengend Snippet: IHC detection of virus antigen expression in mouse tissue after challenge with MERS-CoV. Lung (A–C) and trachea (D–F) sections were assessed using rabbit polyclonal antibody to MERS-CoV nucleoprotein (NP) 3 days after the MERS-CoV challenge. The dark purple spot marked the inflammatory cell infiltration, and the brown particle marked the antigen of MERS-CoV. The MERS-CoV was located mainly in the trachea. Additionally, the lung tissue showed MERS-CoV expression in all immunized groups.

    Article Snippet: Rabbit-serum-derived polyclonal antibody against nucleoprotein (cat: 100213-RP02; Sino Biological Inc., Beijing, CHN) was incubated with the sections at 1:1000 dilution; goat anti-rabbit (cat: pv-9001; ZSGB-Bio, Beijing, CHN) secondary antibody was used at 1:2000, and sections were evaluated using light microscopy.

    Techniques: Immunohistochemistry, Expressing

    rNTD or rRBD vaccination reduced respiratory tract pathology in mice after MERS-CoV challenge. Representative results of hematoxylin-eosin (HE) staining in the lung (A – C) and trachea (D – F) of mock-treated or immunized mice. Severe lesions including the loss of pulmonary alveolus (represented by the white vacuole) and diffuse inflammatory cell infiltration (represented by the dark purple point) are shown (figure A). In contrast, milder lesions were observed among mice immunized with rRBD (figure B) or NTD (figure C), as the pulmonary alveolus was highly visible with less inflammatory cell infiltration. Inflammatory cell infiltration and impaired epithelium of the tunica mucosa bronchiorum were seen in the mock group (D). rRBD (E) or rNTD (F) alleviated the pathologic damage in the trachea of immunized mice. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

    Journal: Vaccine

    Article Title: The recombinant N-terminal domain of spike proteins is a potential vaccine against Middle East respiratory syndrome coronavirus (MERS-CoV) infection

    doi: 10.1016/j.vaccine.2016.11.064

    Figure Lengend Snippet: rNTD or rRBD vaccination reduced respiratory tract pathology in mice after MERS-CoV challenge. Representative results of hematoxylin-eosin (HE) staining in the lung (A – C) and trachea (D – F) of mock-treated or immunized mice. Severe lesions including the loss of pulmonary alveolus (represented by the white vacuole) and diffuse inflammatory cell infiltration (represented by the dark purple point) are shown (figure A). In contrast, milder lesions were observed among mice immunized with rRBD (figure B) or NTD (figure C), as the pulmonary alveolus was highly visible with less inflammatory cell infiltration. Inflammatory cell infiltration and impaired epithelium of the tunica mucosa bronchiorum were seen in the mock group (D). rRBD (E) or rNTD (F) alleviated the pathologic damage in the trachea of immunized mice. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

    Article Snippet: Rabbit-serum-derived polyclonal antibody against nucleoprotein (cat: 100213-RP02; Sino Biological Inc., Beijing, CHN) was incubated with the sections at 1:1000 dilution; goat anti-rabbit (cat: pv-9001; ZSGB-Bio, Beijing, CHN) secondary antibody was used at 1:2000, and sections were evaluated using light microscopy.

    Techniques: Mouse Assay, Staining

    Description of the N-terminal domain (NTD) immunogen and vaccination schedule. (A) The location of the NTD protein on the Middle East respiratory syndrome coronavirus MERS-CoV spike (S) protein. The recombinant (r)NTD protein consists of 336 amino acid (aa) residues (18–353) of S protein. A gp67 signal peptide (SP) was added to the N terminus for expression of the rNTD protein. (B) Purified rNTD protein detected by SDS-PAGE (left) and Western blot (right). The purified rNTD protein was separated by a 10% SDS-PAGE and stained with 0.25% Coomassie brilliant blue. Anti-NTD polyclonal antibody and infrared ray-labeled secondary antibody were used for the Western blot assay. Lane 1: protein molecular weight marker; lane 2: purified rNTD protein. (C). Vaccination schedule and detection. Mice received three vaccinations consisting of 5 or 10 μg of rNTD protein combined with adjuvants at 4-week intervals. Sera were collected at the indicated times to analyze the humoral immune response. Six mice from each group were sacrificed 2 weeks after the last immunization. The spleens were harvested for enzyme-linked immunospot (ELISpot), intracellular cytokine staining (ICS), and cytometric bead array (CBA) assays. In parallel experiments, the remaining mice were challenged with MERS-CoV to detect the protective effect elicited by the rNTD protein. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

    Journal: Vaccine

    Article Title: The recombinant N-terminal domain of spike proteins is a potential vaccine against Middle East respiratory syndrome coronavirus (MERS-CoV) infection

    doi: 10.1016/j.vaccine.2016.11.064

    Figure Lengend Snippet: Description of the N-terminal domain (NTD) immunogen and vaccination schedule. (A) The location of the NTD protein on the Middle East respiratory syndrome coronavirus MERS-CoV spike (S) protein. The recombinant (r)NTD protein consists of 336 amino acid (aa) residues (18–353) of S protein. A gp67 signal peptide (SP) was added to the N terminus for expression of the rNTD protein. (B) Purified rNTD protein detected by SDS-PAGE (left) and Western blot (right). The purified rNTD protein was separated by a 10% SDS-PAGE and stained with 0.25% Coomassie brilliant blue. Anti-NTD polyclonal antibody and infrared ray-labeled secondary antibody were used for the Western blot assay. Lane 1: protein molecular weight marker; lane 2: purified rNTD protein. (C). Vaccination schedule and detection. Mice received three vaccinations consisting of 5 or 10 μg of rNTD protein combined with adjuvants at 4-week intervals. Sera were collected at the indicated times to analyze the humoral immune response. Six mice from each group were sacrificed 2 weeks after the last immunization. The spleens were harvested for enzyme-linked immunospot (ELISpot), intracellular cytokine staining (ICS), and cytometric bead array (CBA) assays. In parallel experiments, the remaining mice were challenged with MERS-CoV to detect the protective effect elicited by the rNTD protein. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

    Article Snippet: Rabbit-serum-derived polyclonal antibody against nucleoprotein (cat: 100213-RP02; Sino Biological Inc., Beijing, CHN) was incubated with the sections at 1:1000 dilution; goat anti-rabbit (cat: pv-9001; ZSGB-Bio, Beijing, CHN) secondary antibody was used at 1:2000, and sections were evaluated using light microscopy.

    Techniques: Recombinant, Expressing, Purification, SDS Page, Western Blot, Staining, Labeling, Molecular Weight, Marker, Mouse Assay, Enzyme-linked Immunospot, Crocin Bleaching Assay

    Pseudo-particle assays of MERS-CoV S WT and mutants. Huh-7 cells were infected with MLV-based pseudo-particles (PP) carrying MERS-CoV S WT or one of the respective S mutants. After 72 h, infected cells were lysed and assessed for luciferase activity. (A) PP infectivity of Huh 7 cells. (B) Infectivity of PP carrying the D922A S protein. Δenv and VSV-G served as representative controls for all PP assays (C) Impact of intracellular Ca 2+ on MERS-CoV fusion. Cells were pre-treated with growth medium containing either 50 µM calcium chelator BAPTA-AM ordimethyl sulfoxide (DMSO) for 1 h. Cells were then infected with their respective PP in the presence of BAPTA-AM or DMSO for 2 h and grown for 72 h before assessing for luciferase activity. (D) Impact of extracellular Ca 2+ on MERS-CoV fusion. Cells were pre-treated with growth medium either with or without 1.8 mM Ca 2+ for 1 h. Infection protocol is as described above except PP were treated with 1.5 mM EGTA for calcium chelation. Infectivity was normalized such that WT PP infectivity is 1. Error bars represent standard deviation (n = 3). Statistical analysis was performed using an unpaired student’s t-test, as indicated. * = p > 0.5, ** = p > 0.05, *** = p > 0.005.

    Journal: bioRxiv

    Article Title: Ca2+ ions promote fusion of Middle East Respiratory Syndrome coronavirus with host cells and increase infectivity

    doi: 10.1101/2019.12.18.881391

    Figure Lengend Snippet: Pseudo-particle assays of MERS-CoV S WT and mutants. Huh-7 cells were infected with MLV-based pseudo-particles (PP) carrying MERS-CoV S WT or one of the respective S mutants. After 72 h, infected cells were lysed and assessed for luciferase activity. (A) PP infectivity of Huh 7 cells. (B) Infectivity of PP carrying the D922A S protein. Δenv and VSV-G served as representative controls for all PP assays (C) Impact of intracellular Ca 2+ on MERS-CoV fusion. Cells were pre-treated with growth medium containing either 50 µM calcium chelator BAPTA-AM ordimethyl sulfoxide (DMSO) for 1 h. Cells were then infected with their respective PP in the presence of BAPTA-AM or DMSO for 2 h and grown for 72 h before assessing for luciferase activity. (D) Impact of extracellular Ca 2+ on MERS-CoV fusion. Cells were pre-treated with growth medium either with or without 1.8 mM Ca 2+ for 1 h. Infection protocol is as described above except PP were treated with 1.5 mM EGTA for calcium chelation. Infectivity was normalized such that WT PP infectivity is 1. Error bars represent standard deviation (n = 3). Statistical analysis was performed using an unpaired student’s t-test, as indicated. * = p > 0.5, ** = p > 0.05, *** = p > 0.005.

    Article Snippet: S protein was detected using the MERS-CoV S rabbit polyclonal antibody (Sino Biological, Cat No: 40069-RP01) as the primary antibody, and AlexaFluor 488-labeled anti-rabbit secondary antibody (Invitrogen).

    Techniques: Infection, Luciferase, Activity Assay, Standard Deviation

    ESR and ITC analysis of the MERS-CoV FP. A-B. Plots of order parameters of DPPTC (A), and 5PC (B) versus peptide:lipid ratio (P/L ratio) of MERS FP or SARS FP in POPC/POPS/Chol=3/1/1 MLVs in buffer with 150 mM NaCl at 25°C. Black, MERS FP, 1 mM Ca 2+ and at pH 5; red, MERS FP calcium-less buffer with 1 mM EGTA and at pH 5; blue, SARS FP, 1 mM Ca 2+ at pH 5, and purple, scrambled peptide, 1 mM Ca 2+ and at pH 5. (C) Plot of difference of order parameters of DPPTC with and without 1% peptide binding (ΔS0) versus Ca 2+ concentration in POPC/POPS/Chol=3/1/1 MLVs in buffer with 150 mM NaCl at 25°C. Black, MERS FP; blue, SARS FP; and green, scrambled peptide. The experiments were typically repeated two to three times. The typical uncertainties found for S 0 ranges from 1-5 × 10 −3 , while the uncertainties from repeated experiments were 5-8 × 10 −3 or less than ±0.01. We show the standard deviation bars in Panel A and B. (D) ITC analysis of Ca 2+ binding to MERS-CoV FP. The peptides were titrated with CaCl 2 . The integrated data represent the enthalpy change per mole of injectant, ΔH, in units of kJ/mol as a function of the molar ratio. Data points and fitted data are overlaid. The fitting is based on the one-site model.

    Journal: bioRxiv

    Article Title: Ca2+ ions promote fusion of Middle East Respiratory Syndrome coronavirus with host cells and increase infectivity

    doi: 10.1101/2019.12.18.881391

    Figure Lengend Snippet: ESR and ITC analysis of the MERS-CoV FP. A-B. Plots of order parameters of DPPTC (A), and 5PC (B) versus peptide:lipid ratio (P/L ratio) of MERS FP or SARS FP in POPC/POPS/Chol=3/1/1 MLVs in buffer with 150 mM NaCl at 25°C. Black, MERS FP, 1 mM Ca 2+ and at pH 5; red, MERS FP calcium-less buffer with 1 mM EGTA and at pH 5; blue, SARS FP, 1 mM Ca 2+ at pH 5, and purple, scrambled peptide, 1 mM Ca 2+ and at pH 5. (C) Plot of difference of order parameters of DPPTC with and without 1% peptide binding (ΔS0) versus Ca 2+ concentration in POPC/POPS/Chol=3/1/1 MLVs in buffer with 150 mM NaCl at 25°C. Black, MERS FP; blue, SARS FP; and green, scrambled peptide. The experiments were typically repeated two to three times. The typical uncertainties found for S 0 ranges from 1-5 × 10 −3 , while the uncertainties from repeated experiments were 5-8 × 10 −3 or less than ±0.01. We show the standard deviation bars in Panel A and B. (D) ITC analysis of Ca 2+ binding to MERS-CoV FP. The peptides were titrated with CaCl 2 . The integrated data represent the enthalpy change per mole of injectant, ΔH, in units of kJ/mol as a function of the molar ratio. Data points and fitted data are overlaid. The fitting is based on the one-site model.

    Article Snippet: S protein was detected using the MERS-CoV S rabbit polyclonal antibody (Sino Biological, Cat No: 40069-RP01) as the primary antibody, and AlexaFluor 488-labeled anti-rabbit secondary antibody (Invitrogen).

    Techniques: Binding Assay, Concentration Assay, Standard Deviation

    Sequence and model of MERS-CoV S fusion loop. (A) Sequences of SARS-CoV S Urbani and MERS-CoV S EMC/2012 fusion peptides (FP). FP1 and FP2 designate the two different domains in the FP as described previously (Lai Millet). Sequences below illustrate the mutations that were introduced in the MERS-CoV S protein via site-directed mutagenesis. In red are the negatively charged residues D and E, in green are the A substitutions. (B) Modeling of the MERS-CoV S monomer with an emphasis on the FP. Negatively charges Ds and E are depicted as atomic bonds in red. The S2’ site is orange and the FP1 and FP2 domains are labeled blue and pink, respectively.

    Journal: bioRxiv

    Article Title: Ca2+ ions promote fusion of Middle East Respiratory Syndrome coronavirus with host cells and increase infectivity

    doi: 10.1101/2019.12.18.881391

    Figure Lengend Snippet: Sequence and model of MERS-CoV S fusion loop. (A) Sequences of SARS-CoV S Urbani and MERS-CoV S EMC/2012 fusion peptides (FP). FP1 and FP2 designate the two different domains in the FP as described previously (Lai Millet). Sequences below illustrate the mutations that were introduced in the MERS-CoV S protein via site-directed mutagenesis. In red are the negatively charged residues D and E, in green are the A substitutions. (B) Modeling of the MERS-CoV S monomer with an emphasis on the FP. Negatively charges Ds and E are depicted as atomic bonds in red. The S2’ site is orange and the FP1 and FP2 domains are labeled blue and pink, respectively.

    Article Snippet: S protein was detected using the MERS-CoV S rabbit polyclonal antibody (Sino Biological, Cat No: 40069-RP01) as the primary antibody, and AlexaFluor 488-labeled anti-rabbit secondary antibody (Invitrogen).

    Techniques: Sequencing, Mutagenesis, Labeling

    Western blot analysis of S proteins incorporated into PPs. 1 ml of DMEM containing PPs per each tested S protein were ultra-centrifuged, washed in PBS and resuspended in SDS Laemmli Buffer. Incorporated S proteins were analyzed using SDS-PAGE and detected using a Western blot with MERS-CoV S antibodies.

    Journal: bioRxiv

    Article Title: Ca2+ ions promote fusion of Middle East Respiratory Syndrome coronavirus with host cells and increase infectivity

    doi: 10.1101/2019.12.18.881391

    Figure Lengend Snippet: Western blot analysis of S proteins incorporated into PPs. 1 ml of DMEM containing PPs per each tested S protein were ultra-centrifuged, washed in PBS and resuspended in SDS Laemmli Buffer. Incorporated S proteins were analyzed using SDS-PAGE and detected using a Western blot with MERS-CoV S antibodies.

    Article Snippet: S protein was detected using the MERS-CoV S rabbit polyclonal antibody (Sino Biological, Cat No: 40069-RP01) as the primary antibody, and AlexaFluor 488-labeled anti-rabbit secondary antibody (Invitrogen).

    Techniques: Western Blot, SDS Page

    Pseudo-particle assays of MERS-CoV S WT and E891A/D896A, E891A/D902A and E891A/D896A/D902A mutants. Huh-7 cells were infected with MLV-based pseudo-particles (PP) carrying MERS-CoV S WT or one of the respective mutants. Infectivity was normalized to WT sample. Error bars represent standard deviation (n = 3). Statistical analysis was performed using an unpaired student’s t-test comparing the WT against the respective mutant (for B and C the untreated WT was compared to each sample). * = p > 0.5, ** = p > 0.05, *** = p > 0.005. (A) Infectivity of PPs without pre-treatment of cells. (B) Impact of intracellular Ca 2+ on MERS-CoV fusion. Cells and PPs were treated as described for Figure 5 C . (C) Impact of extracellular Ca 2+ on MERS-CoV fusion. Cells and PPs were treated as described for Figure 5 D .

    Journal: bioRxiv

    Article Title: Ca2+ ions promote fusion of Middle East Respiratory Syndrome coronavirus with host cells and increase infectivity

    doi: 10.1101/2019.12.18.881391

    Figure Lengend Snippet: Pseudo-particle assays of MERS-CoV S WT and E891A/D896A, E891A/D902A and E891A/D896A/D902A mutants. Huh-7 cells were infected with MLV-based pseudo-particles (PP) carrying MERS-CoV S WT or one of the respective mutants. Infectivity was normalized to WT sample. Error bars represent standard deviation (n = 3). Statistical analysis was performed using an unpaired student’s t-test comparing the WT against the respective mutant (for B and C the untreated WT was compared to each sample). * = p > 0.5, ** = p > 0.05, *** = p > 0.005. (A) Infectivity of PPs without pre-treatment of cells. (B) Impact of intracellular Ca 2+ on MERS-CoV fusion. Cells and PPs were treated as described for Figure 5 C . (C) Impact of extracellular Ca 2+ on MERS-CoV fusion. Cells and PPs were treated as described for Figure 5 D .

    Article Snippet: S protein was detected using the MERS-CoV S rabbit polyclonal antibody (Sino Biological, Cat No: 40069-RP01) as the primary antibody, and AlexaFluor 488-labeled anti-rabbit secondary antibody (Invitrogen).

    Techniques: Infection, Standard Deviation, Mutagenesis

    Immunofluorescence assay of MERS-CoV S WT and mutants. (A) Vero cells were transfected with plasmid DNA encoding for the respective MERS-CoV S variants and the DPP4 binding receptor and grown for 18 h. As Vero cells express endogenous proteases, which cleaves MERS-CoV S for fusion, no further protease treatment was needed to induce syncytia formation. WT + protease inhibitor indicates the condition in which protease inhibitor dec-RVKR-CMK at a concentration of 75 µM was added at the time of transfection to block fusion. Syncytia was visualized using immunofluorescence microscopy by staining the MERS-CoV S with a polyclonal anti-S antibody (in green) and the nuclei with 4′,6-diamidino-2-phenylindole (DAPI, in blue). Images were taken at a magnification of 25x. (B) Quantification of syncytia. Nuclei of 9 syncytia were counted and the average number of nuclei per syncytia was calculated. Error bars represent standard deviation (n = 9). Statistical analysis was performed using an unpaired student’s t-test comparing the WT against each of the respective mutant * = p > 0.5, ** = p > 0.05, *** = p > 0.005.

    Journal: bioRxiv

    Article Title: Ca2+ ions promote fusion of Middle East Respiratory Syndrome coronavirus with host cells and increase infectivity

    doi: 10.1101/2019.12.18.881391

    Figure Lengend Snippet: Immunofluorescence assay of MERS-CoV S WT and mutants. (A) Vero cells were transfected with plasmid DNA encoding for the respective MERS-CoV S variants and the DPP4 binding receptor and grown for 18 h. As Vero cells express endogenous proteases, which cleaves MERS-CoV S for fusion, no further protease treatment was needed to induce syncytia formation. WT + protease inhibitor indicates the condition in which protease inhibitor dec-RVKR-CMK at a concentration of 75 µM was added at the time of transfection to block fusion. Syncytia was visualized using immunofluorescence microscopy by staining the MERS-CoV S with a polyclonal anti-S antibody (in green) and the nuclei with 4′,6-diamidino-2-phenylindole (DAPI, in blue). Images were taken at a magnification of 25x. (B) Quantification of syncytia. Nuclei of 9 syncytia were counted and the average number of nuclei per syncytia was calculated. Error bars represent standard deviation (n = 9). Statistical analysis was performed using an unpaired student’s t-test comparing the WT against each of the respective mutant * = p > 0.5, ** = p > 0.05, *** = p > 0.005.

    Article Snippet: S protein was detected using the MERS-CoV S rabbit polyclonal antibody (Sino Biological, Cat No: 40069-RP01) as the primary antibody, and AlexaFluor 488-labeled anti-rabbit secondary antibody (Invitrogen).

    Techniques: Immunofluorescence, Transfection, Plasmid Preparation, Binding Assay, Protease Inhibitor, Concentration Assay, Blocking Assay, Microscopy, Staining, Standard Deviation, Mutagenesis

    Protein expression and trypsin-mediated cleavage of MERS-CoV S WT and mutants. (A) Plasmid DNA encoding MERS-CoV S WT EMC/2012 was transfected in HEK293T cells. The protease inhibitor dec-RVKR-CMK at a concentration of 75 µM was added at the time of transfection, as indicated. After 18 h, transfected cells were treated with 0.8 nM TPCK-treated trypsin, as indicated. Proteins were subsequently isolated via cell-surface biotinylation. The cell surface proteins were analyzed using SDS-PAGE and detected using a Western blot with MERS-CoV S antibodies. (B) and (C) MERS-CoV S mutant proteins with indicated A substitutions were expressed in HEK293T cells. Protease inhibitor dec-RVKR-CMK was added at the time of transfection and after 18 h, cells were treated with TPCK-treated trypsin, as indicated. Cell surface proteins were isolated and analyzed as described above. Full length S proteins are visible at approx. 250 kDa. S1/S2 cleaved S protein are visible at approx. 115 kDa.

    Journal: bioRxiv

    Article Title: Ca2+ ions promote fusion of Middle East Respiratory Syndrome coronavirus with host cells and increase infectivity

    doi: 10.1101/2019.12.18.881391

    Figure Lengend Snippet: Protein expression and trypsin-mediated cleavage of MERS-CoV S WT and mutants. (A) Plasmid DNA encoding MERS-CoV S WT EMC/2012 was transfected in HEK293T cells. The protease inhibitor dec-RVKR-CMK at a concentration of 75 µM was added at the time of transfection, as indicated. After 18 h, transfected cells were treated with 0.8 nM TPCK-treated trypsin, as indicated. Proteins were subsequently isolated via cell-surface biotinylation. The cell surface proteins were analyzed using SDS-PAGE and detected using a Western blot with MERS-CoV S antibodies. (B) and (C) MERS-CoV S mutant proteins with indicated A substitutions were expressed in HEK293T cells. Protease inhibitor dec-RVKR-CMK was added at the time of transfection and after 18 h, cells were treated with TPCK-treated trypsin, as indicated. Cell surface proteins were isolated and analyzed as described above. Full length S proteins are visible at approx. 250 kDa. S1/S2 cleaved S protein are visible at approx. 115 kDa.

    Article Snippet: S protein was detected using the MERS-CoV S rabbit polyclonal antibody (Sino Biological, Cat No: 40069-RP01) as the primary antibody, and AlexaFluor 488-labeled anti-rabbit secondary antibody (Invitrogen).

    Techniques: Expressing, Plasmid Preparation, Transfection, Protease Inhibitor, Concentration Assay, Isolation, SDS Page, Western Blot, Mutagenesis