sars cov 2 2019 ncov spike orf mammalian expression plasmid codon optimized covid 19 spike research  (Sino Biological)


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    Name:
    SARS CoV 2 2019 nCoV Spike ORF mammalian expression plasmid Codon Optimized COVID 19 Spike Research
    Description:
    Full length Clone DNA of SARS CoV 2 2019 nCoV Spike
    Catalog Number:
    VG40589-UT
    Price:
    670.0
    Category:
    cDNA Clone
    Applications:
    Stable or Transient mammalian expression
    Size:
    1Unit
    Product Aliases:
    coronavirus spike cDNA ORF Clone 2019-nCoV, cov spike cDNA ORF Clone 2019-nCoV, ncov RBD cDNA ORF Clone 2019-nCoV, ncov s1 cDNA ORF Clone 2019-nCoV, ncov s2 cDNA ORF Clone 2019-nCoV, ncov spike cDNA ORF Clone 2019-nCoV, NCP-CoV RBD cDNA ORF Clone 2019-nCoV, NCP-CoV s1 cDNA ORF Clone 2019-nCoV, NCP-CoV s2 cDNA ORF Clone 2019-nCoV, NCP-CoV Spike cDNA ORF Clone 2019-nCoV, novel coronavirus RBD cDNA ORF Clone 2019-nCoV, novel coronavirus s1 cDNA ORF Clone 2019-nCoV, novel coronavirus s2 cDNA ORF Clone 2019-nCoV, novel coronavirus spike cDNA ORF Clone 2019-nCoV, RBD cDNA ORF Clone 2019-nCoV, S1 cDNA ORF Clone 2019-nCoV, S2 cDNA ORF Clone 2019-nCoV, Spike RBD cDNA ORF Clone 2019-nCoV
    Molecule Name:
    Spike
    Buy from Supplier


    Structured Review

    Sino Biological sars cov 2 2019 ncov spike orf mammalian expression plasmid codon optimized covid 19 spike research
    SARS CoV 2 2019 nCoV Spike ORF mammalian expression plasmid Codon Optimized COVID 19 Spike Research
    Full length Clone DNA of SARS CoV 2 2019 nCoV Spike
    https://www.bioz.com/result/sars cov 2 2019 ncov spike orf mammalian expression plasmid codon optimized covid 19 spike research/product/Sino Biological
    Average 98 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    sars cov 2 2019 ncov spike orf mammalian expression plasmid codon optimized covid 19 spike research - by Bioz Stars, 2021-04
    98/100 stars

    Images

    1) Product Images from "Real-Time Conformational Dynamics of SARS-CoV-2 Spikes on Virus Particles"

    Article Title: Real-Time Conformational Dynamics of SARS-CoV-2 Spikes on Virus Particles

    Journal: Cell Host & Microbe

    doi: 10.1016/j.chom.2020.11.001

    Conformational Effects of Trypsin Treatment of Spikes Follow the hACE2-Dependent Activation Pathway (A–C) The serine protease trypsin remodels conformational landscape of spike proteins toward down-stream conformations on the path of hACE2-dependent activation. (A and B) The FRET histogram (A) and TDP (B) of spike proteins on HIV-1 lentivirus particles in the presence of 50 μg/mL trypsin. (C) An experiment as in (A), for spikes in the presence of both 50 μg/mL trypsin and 200 μg/mL hACE2. (D) Three-dimensional presentations of FRET histograms of spike proteins on the virus in the presence and the absence of hACE2 and trypsin. FRET histograms represent mean ± SEM, determined from three randomly assigned populations of FRET traces. For evaluated state occupancies, see Table S1 . (E and F) Trypsin enhances SARS-CoV-2 spike-mediated hACE2-dependent virus-cell fusion. (E) Assay design to monitor virus-cell fusion using the HiBit and LgBiT split NanoLuc system ( Yamamoto et al., 2019 ). Vpr-HiBit was packaged into lentiviral particles carrying SARS-CoV-2 spike (LV_Spike). HEK293 target cells transiently expressing LgBiT tagged to PH domain of human phospholipase Cδ at the N terminus alone or together with hACE2. hACE2-dependent virus-cell fusion was determined by monitoring reconstituted NanoLuc activity in target cells 24 h after infection. (F) Normalized relative luciferase units (RLU; mean ± SD, two replicates with quadruplicates) measured 24 h post-infection to quantify virus-cell fusion in stated target cells after treating viruses with or with indicated amounts of trypsin for 15–20 min at 37°C. NanoLuc activities were normalized to luciferase activity detected in uninfected target cells. p values derived from unpaired t test; ∗∗∗∗ corresponds to p
    Figure Legend Snippet: Conformational Effects of Trypsin Treatment of Spikes Follow the hACE2-Dependent Activation Pathway (A–C) The serine protease trypsin remodels conformational landscape of spike proteins toward down-stream conformations on the path of hACE2-dependent activation. (A and B) The FRET histogram (A) and TDP (B) of spike proteins on HIV-1 lentivirus particles in the presence of 50 μg/mL trypsin. (C) An experiment as in (A), for spikes in the presence of both 50 μg/mL trypsin and 200 μg/mL hACE2. (D) Three-dimensional presentations of FRET histograms of spike proteins on the virus in the presence and the absence of hACE2 and trypsin. FRET histograms represent mean ± SEM, determined from three randomly assigned populations of FRET traces. For evaluated state occupancies, see Table S1 . (E and F) Trypsin enhances SARS-CoV-2 spike-mediated hACE2-dependent virus-cell fusion. (E) Assay design to monitor virus-cell fusion using the HiBit and LgBiT split NanoLuc system ( Yamamoto et al., 2019 ). Vpr-HiBit was packaged into lentiviral particles carrying SARS-CoV-2 spike (LV_Spike). HEK293 target cells transiently expressing LgBiT tagged to PH domain of human phospholipase Cδ at the N terminus alone or together with hACE2. hACE2-dependent virus-cell fusion was determined by monitoring reconstituted NanoLuc activity in target cells 24 h after infection. (F) Normalized relative luciferase units (RLU; mean ± SD, two replicates with quadruplicates) measured 24 h post-infection to quantify virus-cell fusion in stated target cells after treating viruses with or with indicated amounts of trypsin for 15–20 min at 37°C. NanoLuc activities were normalized to luciferase activity detected in uninfected target cells. p values derived from unpaired t test; ∗∗∗∗ corresponds to p

    Techniques Used: Activation Assay, Expressing, Activity Assay, Infection, Luciferase, Derivative Assay

    Related Articles

    Transfection:

    Article Title: An antibody-dependent enhancement (ADE) activity eliminated neutralizing antibody with potent prophylactic and therapeutic efficacy against SARS-CoV-2 in rhesus monkeys
    Article Snippet: Flow cytometry assay The binding of MW05 and MW07 to S protein expression on cell surface was assessed by FACS. .. HEK293 cells were transiently transfected by SARS-CoV-2 Spike expression plasmid (Cat: VG40589-UT, Sino Biological) for 24 to 48 hours. ..

    Article Title: SARS-CoV-2 and SARS-CoV Spike-RBD Structure and Receptor Binding Comparison and Potential Implications on Neutralizing Antibody and Vaccine Development
    Article Snippet: .. Reagents, recombinant proteins and antibodiesRecombinant S1 proteins of SARS-CoV-2 (Cat: 40591-V08H), SARS-CoV (Cat: 40150-V08B1) and MERS-CoV (Cat:40069-V08H), recombinant RBD protein of SARS-CoV (Cat: 40150-V31B2), transfection reagent Sinofection (Cat: STF02), mammalian expression plasmids of full length S or RBD protein of SARS-CoV-2 (Cat: VG40589-UT, Wuhan/IVDC-HB-01/2019) and SARS-CoV (Cat: VG40150-G-N, CUHK-W1), ACE2 (Cat: HG10108-UT), polyclonal antibodies against SARS-CoV RP01 (Cat: 40150-RP01) and T52 (Cat: 40150-T52) were purchased from Sino Biological. ..

    Article Title: Development of a multi-antigenic SARS-CoV-2 vaccine candidate using a synthetic poxvirus platform
    Article Snippet: SARS-CoV-2 pseudovirus production The day before transfection, HEK293T/17 were seeded in a 15-cm dish at a density of 5 × 106 cells in DMEM supplemented with 10% heat inactivated FBS, non-essential amino acids, HEPES, and glutamine . .. Next day, cells were transfected with a mix of packaging vector (pALDI-Lenti System, Aldevron), luciferase reporter vector, and a plasmid encoding for the wild-type SARS-CoV2 Spike protein (VG40589-UT, Sino Biological) or vesicular stomatitis virus G (VSV-G, Aldevron), using FuGENE6 (Roche) as a transfection reagent:DNA ratio of 3:1, according to manufacturer’s protocol. .. Sixteen hours post-transfection, the media was replaced and cells were incubated for an additional 24–72 h. Supernatants were harvested at 24, 48, and 72 h, clarified by centrifugation at 1500 r.p.m. for 5 min and filtered using a sterile 0.22-µm pore size filter.

    Expressing:

    Article Title: An antibody-dependent enhancement (ADE) activity eliminated neutralizing antibody with potent prophylactic and therapeutic efficacy against SARS-CoV-2 in rhesus monkeys
    Article Snippet: Flow cytometry assay The binding of MW05 and MW07 to S protein expression on cell surface was assessed by FACS. .. HEK293 cells were transiently transfected by SARS-CoV-2 Spike expression plasmid (Cat: VG40589-UT, Sino Biological) for 24 to 48 hours. ..

    Article Title: Naturally mutated spike proteins of SARS-CoV-2 variants show differential levels of cell entry
    Article Snippet: The SARS-CoV S expression plasmid pC-SARS-S was created by inserting the BsiWI/XhoI-digested PCR-amplified SARS-CoV S fragment of CMV/R-SARS-S into the corresponding site of pCAGGS. .. The SARS-CoV-2 S expression plasmid pC-SARS2-S was created by inserting the Acc65I/NotI-digested PCR-amplified SARS-CoV-2 S fragment of pCMV3-2019-nCoV-Spike(S1+S2)-long (Sino Biological; VG40589-UT) into the corresponding site of pCAGGS. .. The SARS-CoV-2 S mutants (pC-SARS2-S-H49Y, pC-SARS2-S-V367F, pC-SARS2-S-G476S, pC-SARS2-S-V483A, pC-SARS2-S-D614G, or pC-SARS2-S-C1247A), in which positions 49, 367, 476, 483, 614, or 1247 of the S protein were mutated from histidine to tyrosine, valine to phenylalanine, glycine to serine, valine to alanine, aspartic acid to glycine, or cysteine to alanine, respectively, were created by inserting overlapping PCR fragments into Acc65I/NotI-digested pCAGGS.

    Article Title: SARS-CoV-2 and SARS-CoV Spike-RBD Structure and Receptor Binding Comparison and Potential Implications on Neutralizing Antibody and Vaccine Development
    Article Snippet: .. Reagents, recombinant proteins and antibodiesRecombinant S1 proteins of SARS-CoV-2 (Cat: 40591-V08H), SARS-CoV (Cat: 40150-V08B1) and MERS-CoV (Cat:40069-V08H), recombinant RBD protein of SARS-CoV (Cat: 40150-V31B2), transfection reagent Sinofection (Cat: STF02), mammalian expression plasmids of full length S or RBD protein of SARS-CoV-2 (Cat: VG40589-UT, Wuhan/IVDC-HB-01/2019) and SARS-CoV (Cat: VG40150-G-N, CUHK-W1), ACE2 (Cat: HG10108-UT), polyclonal antibodies against SARS-CoV RP01 (Cat: 40150-RP01) and T52 (Cat: 40150-T52) were purchased from Sino Biological. ..

    Plasmid Preparation:

    Article Title: An antibody-dependent enhancement (ADE) activity eliminated neutralizing antibody with potent prophylactic and therapeutic efficacy against SARS-CoV-2 in rhesus monkeys
    Article Snippet: Flow cytometry assay The binding of MW05 and MW07 to S protein expression on cell surface was assessed by FACS. .. HEK293 cells were transiently transfected by SARS-CoV-2 Spike expression plasmid (Cat: VG40589-UT, Sino Biological) for 24 to 48 hours. ..

    Article Title: Naturally mutated spike proteins of SARS-CoV-2 variants show differential levels of cell entry
    Article Snippet: The SARS-CoV S expression plasmid pC-SARS-S was created by inserting the BsiWI/XhoI-digested PCR-amplified SARS-CoV S fragment of CMV/R-SARS-S into the corresponding site of pCAGGS. .. The SARS-CoV-2 S expression plasmid pC-SARS2-S was created by inserting the Acc65I/NotI-digested PCR-amplified SARS-CoV-2 S fragment of pCMV3-2019-nCoV-Spike(S1+S2)-long (Sino Biological; VG40589-UT) into the corresponding site of pCAGGS. .. The SARS-CoV-2 S mutants (pC-SARS2-S-H49Y, pC-SARS2-S-V367F, pC-SARS2-S-G476S, pC-SARS2-S-V483A, pC-SARS2-S-D614G, or pC-SARS2-S-C1247A), in which positions 49, 367, 476, 483, 614, or 1247 of the S protein were mutated from histidine to tyrosine, valine to phenylalanine, glycine to serine, valine to alanine, aspartic acid to glycine, or cysteine to alanine, respectively, were created by inserting overlapping PCR fragments into Acc65I/NotI-digested pCAGGS.

    Article Title: Development of a multi-antigenic SARS-CoV-2 vaccine candidate using a synthetic poxvirus platform
    Article Snippet: SARS-CoV-2 pseudovirus production The day before transfection, HEK293T/17 were seeded in a 15-cm dish at a density of 5 × 106 cells in DMEM supplemented with 10% heat inactivated FBS, non-essential amino acids, HEPES, and glutamine . .. Next day, cells were transfected with a mix of packaging vector (pALDI-Lenti System, Aldevron), luciferase reporter vector, and a plasmid encoding for the wild-type SARS-CoV2 Spike protein (VG40589-UT, Sino Biological) or vesicular stomatitis virus G (VSV-G, Aldevron), using FuGENE6 (Roche) as a transfection reagent:DNA ratio of 3:1, according to manufacturer’s protocol. .. Sixteen hours post-transfection, the media was replaced and cells were incubated for an additional 24–72 h. Supernatants were harvested at 24, 48, and 72 h, clarified by centrifugation at 1500 r.p.m. for 5 min and filtered using a sterile 0.22-µm pore size filter.

    Article Title: Real-Time Conformational Dynamics of SARS-CoV-2 Spikes on Virus Particles
    Article Snippet: .. Construction of Full-Length Tagged SARS-CoV-2 Spike (S)A full-length wild-type pCMV3-SARS-CoV-2 Spike (S1+S2)-long (termed as pCMV-S, codon-optimized, Sino Biological, cat # VG40589-UT) plasmid was used as a template to generate tagged pCMV-S. .. The translated amino acid sequence of pCMV-S is identical to QHD43416.1 (GenBank).

    Polymerase Chain Reaction:

    Article Title: Naturally mutated spike proteins of SARS-CoV-2 variants show differential levels of cell entry
    Article Snippet: The SARS-CoV S expression plasmid pC-SARS-S was created by inserting the BsiWI/XhoI-digested PCR-amplified SARS-CoV S fragment of CMV/R-SARS-S into the corresponding site of pCAGGS. .. The SARS-CoV-2 S expression plasmid pC-SARS2-S was created by inserting the Acc65I/NotI-digested PCR-amplified SARS-CoV-2 S fragment of pCMV3-2019-nCoV-Spike(S1+S2)-long (Sino Biological; VG40589-UT) into the corresponding site of pCAGGS. .. The SARS-CoV-2 S mutants (pC-SARS2-S-H49Y, pC-SARS2-S-V367F, pC-SARS2-S-G476S, pC-SARS2-S-V483A, pC-SARS2-S-D614G, or pC-SARS2-S-C1247A), in which positions 49, 367, 476, 483, 614, or 1247 of the S protein were mutated from histidine to tyrosine, valine to phenylalanine, glycine to serine, valine to alanine, aspartic acid to glycine, or cysteine to alanine, respectively, were created by inserting overlapping PCR fragments into Acc65I/NotI-digested pCAGGS.

    Recombinant:

    Article Title: SARS-CoV-2 and SARS-CoV Spike-RBD Structure and Receptor Binding Comparison and Potential Implications on Neutralizing Antibody and Vaccine Development
    Article Snippet: .. Reagents, recombinant proteins and antibodiesRecombinant S1 proteins of SARS-CoV-2 (Cat: 40591-V08H), SARS-CoV (Cat: 40150-V08B1) and MERS-CoV (Cat:40069-V08H), recombinant RBD protein of SARS-CoV (Cat: 40150-V31B2), transfection reagent Sinofection (Cat: STF02), mammalian expression plasmids of full length S or RBD protein of SARS-CoV-2 (Cat: VG40589-UT, Wuhan/IVDC-HB-01/2019) and SARS-CoV (Cat: VG40150-G-N, CUHK-W1), ACE2 (Cat: HG10108-UT), polyclonal antibodies against SARS-CoV RP01 (Cat: 40150-RP01) and T52 (Cat: 40150-T52) were purchased from Sino Biological. ..

    Luciferase:

    Article Title: Development of a multi-antigenic SARS-CoV-2 vaccine candidate using a synthetic poxvirus platform
    Article Snippet: SARS-CoV-2 pseudovirus production The day before transfection, HEK293T/17 were seeded in a 15-cm dish at a density of 5 × 106 cells in DMEM supplemented with 10% heat inactivated FBS, non-essential amino acids, HEPES, and glutamine . .. Next day, cells were transfected with a mix of packaging vector (pALDI-Lenti System, Aldevron), luciferase reporter vector, and a plasmid encoding for the wild-type SARS-CoV2 Spike protein (VG40589-UT, Sino Biological) or vesicular stomatitis virus G (VSV-G, Aldevron), using FuGENE6 (Roche) as a transfection reagent:DNA ratio of 3:1, according to manufacturer’s protocol. .. Sixteen hours post-transfection, the media was replaced and cells were incubated for an additional 24–72 h. Supernatants were harvested at 24, 48, and 72 h, clarified by centrifugation at 1500 r.p.m. for 5 min and filtered using a sterile 0.22-µm pore size filter.

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    Sino Biological sars cov 2 2019 ncov spike orf mammalian expression plasmid codon optimized covid 19 spike research
    Conformational Effects of Trypsin Treatment of Spikes Follow the hACE2-Dependent Activation Pathway (A–C) The serine protease trypsin remodels conformational landscape of spike proteins toward down-stream conformations on the path of hACE2-dependent activation. (A and B) The FRET histogram (A) and TDP (B) of spike proteins on HIV-1 lentivirus particles in the presence of 50 μg/mL trypsin. (C) An experiment as in (A), for spikes in the presence of both 50 μg/mL trypsin and 200 μg/mL hACE2. (D) Three-dimensional presentations of FRET histograms of spike proteins on the virus in the presence and the absence of hACE2 and trypsin. FRET histograms represent mean ± SEM, determined from three randomly assigned populations of FRET traces. For evaluated state occupancies, see Table S1 . (E and F) Trypsin enhances <t>SARS-CoV-2</t> spike-mediated hACE2-dependent virus-cell fusion. (E) Assay design to monitor virus-cell fusion using the HiBit and LgBiT split NanoLuc system ( Yamamoto et al., <t>2019</t> ). Vpr-HiBit was packaged into lentiviral particles carrying SARS-CoV-2 spike (LV_Spike). HEK293 target cells transiently expressing LgBiT tagged to PH domain of human phospholipase Cδ at the N terminus alone or together with hACE2. hACE2-dependent virus-cell fusion was determined by monitoring reconstituted NanoLuc activity in target cells 24 h after infection. (F) Normalized relative luciferase units (RLU; mean ± SD, two replicates with quadruplicates) measured 24 h post-infection to quantify virus-cell fusion in stated target cells after treating viruses with or with indicated amounts of trypsin for 15–20 min at 37°C. NanoLuc activities were normalized to luciferase activity detected in uninfected target cells. p values derived from unpaired t test; ∗∗∗∗ corresponds to p
    Sars Cov 2 2019 Ncov Spike Orf Mammalian Expression Plasmid Codon Optimized Covid 19 Spike Research, supplied by Sino Biological, used in various techniques. Bioz Stars score: 98/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/sars cov 2 2019 ncov spike orf mammalian expression plasmid codon optimized covid 19 spike research/product/Sino Biological
    Average 98 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    sars cov 2 2019 ncov spike orf mammalian expression plasmid codon optimized covid 19 spike research - by Bioz Stars, 2021-04
    98/100 stars
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    99
    Sino Biological codon optimized sars cov 2 spike orf mammalian expression plasmid
    W4P-RBD potentiates functional T cells specific to <t>SARS-CoV-2</t> S1 proteins. C57BL/6 mice were intramuscularly injected with W-RBD, W4P-RBD (50 μ g /mouse), or mock, and the spleens were collected 5 weeks post-vaccination for analysis by flow cytometry. (A,B) Splenocytes were incubated with SARS-CoV-2 S1 protein (5 μ g / ml ) for 24 h and stained to detect IFNγ-producing CD8 + T cells and CD4 + T cells. (C) Correlation between RBD-specific IgG in serum and the S1-specific T-cell population in splenocytes. Significance differences (* P
    Codon Optimized Sars Cov 2 Spike Orf Mammalian Expression Plasmid, supplied by Sino Biological, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/codon optimized sars cov 2 spike orf mammalian expression plasmid/product/Sino Biological
    Average 99 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    codon optimized sars cov 2 spike orf mammalian expression plasmid - by Bioz Stars, 2021-04
    99/100 stars
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    93
    Sino Biological sars cov 2 2019 ncov nucleocapsid orf mammalian expression plasmid codon optimized covid 19 nucleocapsid research
    Surface antigenic similarity of rVSV-ΔG-spike and <t>SARS-CoV-2:</t> (A) Immunofluorescent images of Vero E6 cells infected with either WT-VSV (left panel), rVSV-ΔG-spike (middle panel), or SARS-CoV-2, stained with serum from COVID-19 human convalescent serum (right panel). (B) Correlation analysis of neutralization of rVSV-ΔG-spike and SARS-CoV-2 by a panel of sera from COVID-19 convalescent patients. For each sera (n=12), NT 50 values were determined for neutralization of rVSV-ΔG-spike, or SARS-CoV-2. The NT 50 values were plotted to determine the correlation between th neutralization assays. R 2 =0.911.
    Sars Cov 2 2019 Ncov Nucleocapsid Orf Mammalian Expression Plasmid Codon Optimized Covid 19 Nucleocapsid Research, supplied by Sino Biological, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/sars cov 2 2019 ncov nucleocapsid orf mammalian expression plasmid codon optimized covid 19 nucleocapsid research/product/Sino Biological
    Average 93 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    sars cov 2 2019 ncov nucleocapsid orf mammalian expression plasmid codon optimized covid 19 nucleocapsid research - by Bioz Stars, 2021-04
    93/100 stars
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    Conformational Effects of Trypsin Treatment of Spikes Follow the hACE2-Dependent Activation Pathway (A–C) The serine protease trypsin remodels conformational landscape of spike proteins toward down-stream conformations on the path of hACE2-dependent activation. (A and B) The FRET histogram (A) and TDP (B) of spike proteins on HIV-1 lentivirus particles in the presence of 50 μg/mL trypsin. (C) An experiment as in (A), for spikes in the presence of both 50 μg/mL trypsin and 200 μg/mL hACE2. (D) Three-dimensional presentations of FRET histograms of spike proteins on the virus in the presence and the absence of hACE2 and trypsin. FRET histograms represent mean ± SEM, determined from three randomly assigned populations of FRET traces. For evaluated state occupancies, see Table S1 . (E and F) Trypsin enhances SARS-CoV-2 spike-mediated hACE2-dependent virus-cell fusion. (E) Assay design to monitor virus-cell fusion using the HiBit and LgBiT split NanoLuc system ( Yamamoto et al., 2019 ). Vpr-HiBit was packaged into lentiviral particles carrying SARS-CoV-2 spike (LV_Spike). HEK293 target cells transiently expressing LgBiT tagged to PH domain of human phospholipase Cδ at the N terminus alone or together with hACE2. hACE2-dependent virus-cell fusion was determined by monitoring reconstituted NanoLuc activity in target cells 24 h after infection. (F) Normalized relative luciferase units (RLU; mean ± SD, two replicates with quadruplicates) measured 24 h post-infection to quantify virus-cell fusion in stated target cells after treating viruses with or with indicated amounts of trypsin for 15–20 min at 37°C. NanoLuc activities were normalized to luciferase activity detected in uninfected target cells. p values derived from unpaired t test; ∗∗∗∗ corresponds to p

    Journal: Cell Host & Microbe

    Article Title: Real-Time Conformational Dynamics of SARS-CoV-2 Spikes on Virus Particles

    doi: 10.1016/j.chom.2020.11.001

    Figure Lengend Snippet: Conformational Effects of Trypsin Treatment of Spikes Follow the hACE2-Dependent Activation Pathway (A–C) The serine protease trypsin remodels conformational landscape of spike proteins toward down-stream conformations on the path of hACE2-dependent activation. (A and B) The FRET histogram (A) and TDP (B) of spike proteins on HIV-1 lentivirus particles in the presence of 50 μg/mL trypsin. (C) An experiment as in (A), for spikes in the presence of both 50 μg/mL trypsin and 200 μg/mL hACE2. (D) Three-dimensional presentations of FRET histograms of spike proteins on the virus in the presence and the absence of hACE2 and trypsin. FRET histograms represent mean ± SEM, determined from three randomly assigned populations of FRET traces. For evaluated state occupancies, see Table S1 . (E and F) Trypsin enhances SARS-CoV-2 spike-mediated hACE2-dependent virus-cell fusion. (E) Assay design to monitor virus-cell fusion using the HiBit and LgBiT split NanoLuc system ( Yamamoto et al., 2019 ). Vpr-HiBit was packaged into lentiviral particles carrying SARS-CoV-2 spike (LV_Spike). HEK293 target cells transiently expressing LgBiT tagged to PH domain of human phospholipase Cδ at the N terminus alone or together with hACE2. hACE2-dependent virus-cell fusion was determined by monitoring reconstituted NanoLuc activity in target cells 24 h after infection. (F) Normalized relative luciferase units (RLU; mean ± SD, two replicates with quadruplicates) measured 24 h post-infection to quantify virus-cell fusion in stated target cells after treating viruses with or with indicated amounts of trypsin for 15–20 min at 37°C. NanoLuc activities were normalized to luciferase activity detected in uninfected target cells. p values derived from unpaired t test; ∗∗∗∗ corresponds to p

    Article Snippet: Construction of Full-Length Tagged SARS-CoV-2 Spike (S)A full-length wild-type pCMV3-SARS-CoV-2 Spike (S1+S2)-long (termed as pCMV-S, codon-optimized, Sino Biological, cat # VG40589-UT) plasmid was used as a template to generate tagged pCMV-S.

    Techniques: Activation Assay, Expressing, Activity Assay, Infection, Luciferase, Derivative Assay

    Structural conservation of SARS-CoV RBD. RBD is shown as colored surface. ACE2 is shown as gray cartoon. The three surface mutation sites (i.e. N354D, D364Y, and V367F) observed in SARS-CoV-2 RBD are labeled. Mutation F342L is buried and not shown here.

    Journal: bioRxiv

    Article Title: SARS-CoV-2 and SARS-CoV Spike-RBD Structure and Receptor Binding Comparison and Potential Implications on Neutralizing Antibody and Vaccine Development

    doi: 10.1101/2020.02.16.951723

    Figure Lengend Snippet: Structural conservation of SARS-CoV RBD. RBD is shown as colored surface. ACE2 is shown as gray cartoon. The three surface mutation sites (i.e. N354D, D364Y, and V367F) observed in SARS-CoV-2 RBD are labeled. Mutation F342L is buried and not shown here.

    Article Snippet: Reagents, recombinant proteins and antibodies Recombinant S1 proteins of SARS-CoV-2 (Cat: 40591-V08H), SARS-CoV (Cat: 40150-V08B1) and MERS-CoV (Cat:40069-V08H), recombinant RBD protein of SARS-CoV (Cat: 40150-V31B2), transfection reagent Sinofection (Cat: STF02), mammalian expression plasmids of full length S or RBD protein of SARS-CoV-2 (Cat: VG40589-UT, Wuhan/IVDC-HB-01/2019) and SARS-CoV (Cat: VG40150-G-N, CUHK-W1), ACE2 (Cat: HG10108-UT), polyclonal antibodies against SARS-CoV RP01 (Cat: 40150-RP01) and T52 (Cat: 40150-T52) were purchased from Sino Biological.

    Techniques: Mutagenesis, Labeling

    Structure similarity between SARS-CoV-2 RBD and SARS-CoV RBD. RBD is shown in a space-filled model with colored surface. ACE2 is shown as gray tube model. The three glycosylation sites in SARS-CoV are labeled. Note that N 357 ST in SARS-CoV is changed to N 370 SA in SARS-CoV-2, which is different from the NXS/T pattern required for glycosylation, and hence this site is more likely to be unglycosylated. The two possible cross-reactive regions are marked with yellow circles.

    Journal: bioRxiv

    Article Title: SARS-CoV-2 and SARS-CoV Spike-RBD Structure and Receptor Binding Comparison and Potential Implications on Neutralizing Antibody and Vaccine Development

    doi: 10.1101/2020.02.16.951723

    Figure Lengend Snippet: Structure similarity between SARS-CoV-2 RBD and SARS-CoV RBD. RBD is shown in a space-filled model with colored surface. ACE2 is shown as gray tube model. The three glycosylation sites in SARS-CoV are labeled. Note that N 357 ST in SARS-CoV is changed to N 370 SA in SARS-CoV-2, which is different from the NXS/T pattern required for glycosylation, and hence this site is more likely to be unglycosylated. The two possible cross-reactive regions are marked with yellow circles.

    Article Snippet: Reagents, recombinant proteins and antibodies Recombinant S1 proteins of SARS-CoV-2 (Cat: 40591-V08H), SARS-CoV (Cat: 40150-V08B1) and MERS-CoV (Cat:40069-V08H), recombinant RBD protein of SARS-CoV (Cat: 40150-V31B2), transfection reagent Sinofection (Cat: STF02), mammalian expression plasmids of full length S or RBD protein of SARS-CoV-2 (Cat: VG40589-UT, Wuhan/IVDC-HB-01/2019) and SARS-CoV (Cat: VG40150-G-N, CUHK-W1), ACE2 (Cat: HG10108-UT), polyclonal antibodies against SARS-CoV RP01 (Cat: 40150-RP01) and T52 (Cat: 40150-T52) were purchased from Sino Biological.

    Techniques: Labeling

    Cross-reactivity and neutralization efficiency of SARS nAbs against SARS-CoV-2. A. Binding of SARS nAbs to SARS-CoV S1 protein were tested by ELISA. Recombinant S1 protein of SARS-CoV were coated on plates, serial diluted nAbs were added for binding to recombinant S1 protein. B. Binding of SARS nAbs to SARS-CoV-2 S1 protein were tested by ELSIA. Recombinant S1 protein of SARS-CoV-2 were coated on plates, serial diluted nAbs were added for binding to recombinant S1 protein. C. Neutralization of SARS-CoV nAbs against SARS-CoV-2 PSV. D. Antibody competition with SARS-CoV RBD binding to ACE2. Recombinant SARS-CoV RBD protein was coated on plates, nAbs and recombinant ACE2 were then added for RBD binding competition measurements.

    Journal: bioRxiv

    Article Title: SARS-CoV-2 and SARS-CoV Spike-RBD Structure and Receptor Binding Comparison and Potential Implications on Neutralizing Antibody and Vaccine Development

    doi: 10.1101/2020.02.16.951723

    Figure Lengend Snippet: Cross-reactivity and neutralization efficiency of SARS nAbs against SARS-CoV-2. A. Binding of SARS nAbs to SARS-CoV S1 protein were tested by ELISA. Recombinant S1 protein of SARS-CoV were coated on plates, serial diluted nAbs were added for binding to recombinant S1 protein. B. Binding of SARS nAbs to SARS-CoV-2 S1 protein were tested by ELSIA. Recombinant S1 protein of SARS-CoV-2 were coated on plates, serial diluted nAbs were added for binding to recombinant S1 protein. C. Neutralization of SARS-CoV nAbs against SARS-CoV-2 PSV. D. Antibody competition with SARS-CoV RBD binding to ACE2. Recombinant SARS-CoV RBD protein was coated on plates, nAbs and recombinant ACE2 were then added for RBD binding competition measurements.

    Article Snippet: Reagents, recombinant proteins and antibodies Recombinant S1 proteins of SARS-CoV-2 (Cat: 40591-V08H), SARS-CoV (Cat: 40150-V08B1) and MERS-CoV (Cat:40069-V08H), recombinant RBD protein of SARS-CoV (Cat: 40150-V31B2), transfection reagent Sinofection (Cat: STF02), mammalian expression plasmids of full length S or RBD protein of SARS-CoV-2 (Cat: VG40589-UT, Wuhan/IVDC-HB-01/2019) and SARS-CoV (Cat: VG40150-G-N, CUHK-W1), ACE2 (Cat: HG10108-UT), polyclonal antibodies against SARS-CoV RP01 (Cat: 40150-RP01) and T52 (Cat: 40150-T52) were purchased from Sino Biological.

    Techniques: Neutralization, Binding Assay, Enzyme-linked Immunosorbent Assay, Recombinant

    Sequence analysis and structure modeling of SARS-CoV-2 RBD and SARS-CoV RBD and their interactions with ACE2. A. RBD sequence alignment of SARS-CoV and SARS-CoV-2, highlighting the predominant residues that contribute to the interactions with ACE2. The distinct interactions of RBD and ACE2 for the two viruses are indicated by the down-pointing orange triangles and up-pointing red triangles, respectively. RBM residues are underlined. The one-residue insertion is indicated by the red arrow. Asterisks indicate positions of fully conserved residues. Colons indicate positions of strictly conserved residues. Periods indicate positions of weakly conserved residues. B. Conformational comparison between the RBD-ACE2 complex structures for SARS-CoV-2 and SARS-CoV. The RBD and ACE2 structures in the SARS-CoV-2 RBD-ACE2 complex model are shown as orange and pink tubes, respectively. The RBD and ACE2 structures in the optimized SARS-CoV RBD-ACE2 complex structure are shown as blue and green tubes, respectively. The location of noticeable subtle conformational difference is indicated by an arrow. C. Distinct interaction patterns in the SARS-CoV-2 and SARS-CoV RBD-ACE2 interfaces. Structures of RBD and ACE2 are shown as cartoon in pink and green colors, respectively. The side chains of the residues in both protein components, representing their unique interactions, are shown as sticks. Polar interactions (salt-bridge and hydrogen bond) are shown as blue dash line. Non-polar interactions (π-stack, π-anion, and hydrophobic interactions) are shown as orange dash line.

    Journal: bioRxiv

    Article Title: SARS-CoV-2 and SARS-CoV Spike-RBD Structure and Receptor Binding Comparison and Potential Implications on Neutralizing Antibody and Vaccine Development

    doi: 10.1101/2020.02.16.951723

    Figure Lengend Snippet: Sequence analysis and structure modeling of SARS-CoV-2 RBD and SARS-CoV RBD and their interactions with ACE2. A. RBD sequence alignment of SARS-CoV and SARS-CoV-2, highlighting the predominant residues that contribute to the interactions with ACE2. The distinct interactions of RBD and ACE2 for the two viruses are indicated by the down-pointing orange triangles and up-pointing red triangles, respectively. RBM residues are underlined. The one-residue insertion is indicated by the red arrow. Asterisks indicate positions of fully conserved residues. Colons indicate positions of strictly conserved residues. Periods indicate positions of weakly conserved residues. B. Conformational comparison between the RBD-ACE2 complex structures for SARS-CoV-2 and SARS-CoV. The RBD and ACE2 structures in the SARS-CoV-2 RBD-ACE2 complex model are shown as orange and pink tubes, respectively. The RBD and ACE2 structures in the optimized SARS-CoV RBD-ACE2 complex structure are shown as blue and green tubes, respectively. The location of noticeable subtle conformational difference is indicated by an arrow. C. Distinct interaction patterns in the SARS-CoV-2 and SARS-CoV RBD-ACE2 interfaces. Structures of RBD and ACE2 are shown as cartoon in pink and green colors, respectively. The side chains of the residues in both protein components, representing their unique interactions, are shown as sticks. Polar interactions (salt-bridge and hydrogen bond) are shown as blue dash line. Non-polar interactions (π-stack, π-anion, and hydrophobic interactions) are shown as orange dash line.

    Article Snippet: Reagents, recombinant proteins and antibodies Recombinant S1 proteins of SARS-CoV-2 (Cat: 40591-V08H), SARS-CoV (Cat: 40150-V08B1) and MERS-CoV (Cat:40069-V08H), recombinant RBD protein of SARS-CoV (Cat: 40150-V31B2), transfection reagent Sinofection (Cat: STF02), mammalian expression plasmids of full length S or RBD protein of SARS-CoV-2 (Cat: VG40589-UT, Wuhan/IVDC-HB-01/2019) and SARS-CoV (Cat: VG40150-G-N, CUHK-W1), ACE2 (Cat: HG10108-UT), polyclonal antibodies against SARS-CoV RP01 (Cat: 40150-RP01) and T52 (Cat: 40150-T52) were purchased from Sino Biological.

    Techniques: Sequencing

    Measurements of SARS-CoV-2 and SARS-CoV S1 binding to ACE2. A. Serial diluted recombinant S1 proteins of SARS-CoV-2, SARS-CoV and MERS-CoV were coated on 96 well plates, incubated with the recombinant Fc-tagged ACE2 (ACE2-Fc) for binding evaluation. B. Recombinant S1 proteins of SARS-CoV-2 and SARS-CoV were incubated with 293T-ACE2 cells and subjected to FACS evaluation for binding.

    Journal: bioRxiv

    Article Title: SARS-CoV-2 and SARS-CoV Spike-RBD Structure and Receptor Binding Comparison and Potential Implications on Neutralizing Antibody and Vaccine Development

    doi: 10.1101/2020.02.16.951723

    Figure Lengend Snippet: Measurements of SARS-CoV-2 and SARS-CoV S1 binding to ACE2. A. Serial diluted recombinant S1 proteins of SARS-CoV-2, SARS-CoV and MERS-CoV were coated on 96 well plates, incubated with the recombinant Fc-tagged ACE2 (ACE2-Fc) for binding evaluation. B. Recombinant S1 proteins of SARS-CoV-2 and SARS-CoV were incubated with 293T-ACE2 cells and subjected to FACS evaluation for binding.

    Article Snippet: Reagents, recombinant proteins and antibodies Recombinant S1 proteins of SARS-CoV-2 (Cat: 40591-V08H), SARS-CoV (Cat: 40150-V08B1) and MERS-CoV (Cat:40069-V08H), recombinant RBD protein of SARS-CoV (Cat: 40150-V31B2), transfection reagent Sinofection (Cat: STF02), mammalian expression plasmids of full length S or RBD protein of SARS-CoV-2 (Cat: VG40589-UT, Wuhan/IVDC-HB-01/2019) and SARS-CoV (Cat: VG40150-G-N, CUHK-W1), ACE2 (Cat: HG10108-UT), polyclonal antibodies against SARS-CoV RP01 (Cat: 40150-RP01) and T52 (Cat: 40150-T52) were purchased from Sino Biological.

    Techniques: Binding Assay, Recombinant, Incubation, FACS

    W4P-RBD potentiates functional T cells specific to SARS-CoV-2 S1 proteins. C57BL/6 mice were intramuscularly injected with W-RBD, W4P-RBD (50 μ g /mouse), or mock, and the spleens were collected 5 weeks post-vaccination for analysis by flow cytometry. (A,B) Splenocytes were incubated with SARS-CoV-2 S1 protein (5 μ g / ml ) for 24 h and stained to detect IFNγ-producing CD8 + T cells and CD4 + T cells. (C) Correlation between RBD-specific IgG in serum and the S1-specific T-cell population in splenocytes. Significance differences (* P

    Journal: Frontiers in Immunology

    Article Title: A Novel DNA Vaccine Against SARS-CoV-2 Encoding a Chimeric Protein of Its Receptor-Binding Domain (RBD) Fused to the Amino-Terminal Region of Hepatitis B Virus preS1 With a W4P Mutation

    doi: 10.3389/fimmu.2021.637654

    Figure Lengend Snippet: W4P-RBD potentiates functional T cells specific to SARS-CoV-2 S1 proteins. C57BL/6 mice were intramuscularly injected with W-RBD, W4P-RBD (50 μ g /mouse), or mock, and the spleens were collected 5 weeks post-vaccination for analysis by flow cytometry. (A,B) Splenocytes were incubated with SARS-CoV-2 S1 protein (5 μ g / ml ) for 24 h and stained to detect IFNγ-producing CD8 + T cells and CD4 + T cells. (C) Correlation between RBD-specific IgG in serum and the S1-specific T-cell population in splenocytes. Significance differences (* P

    Article Snippet: Briefly, genes encoding RBD (residue 319-541) of the SARS-CoV-2 spike protein were amplified using RBD primers and codon-optimized SARS-CoV-2 Spike ORF mammalian expression plasmid (VG40589; Sino Biological, CN) as the template.

    Techniques: Functional Assay, Mouse Assay, Injection, Flow Cytometry, Incubation, Staining

    Schematic representation of W4P-RBD as a vaccine candidate against SARS-CoV-2. A novel platform of N-terminal addition of HBV W4P preS1 33-bp sequences for a DNA vaccine against SARS-CoV-2 were developed. The W4P-RBD led to enhanced both humoral and cell-mediated immune response against SARS-CoV-2 in vaccinated mice, demonstrating its feasibility as a DNA vaccine to protect against SARS-CoV-2.

    Journal: Frontiers in Immunology

    Article Title: A Novel DNA Vaccine Against SARS-CoV-2 Encoding a Chimeric Protein of Its Receptor-Binding Domain (RBD) Fused to the Amino-Terminal Region of Hepatitis B Virus preS1 With a W4P Mutation

    doi: 10.3389/fimmu.2021.637654

    Figure Lengend Snippet: Schematic representation of W4P-RBD as a vaccine candidate against SARS-CoV-2. A novel platform of N-terminal addition of HBV W4P preS1 33-bp sequences for a DNA vaccine against SARS-CoV-2 were developed. The W4P-RBD led to enhanced both humoral and cell-mediated immune response against SARS-CoV-2 in vaccinated mice, demonstrating its feasibility as a DNA vaccine to protect against SARS-CoV-2.

    Article Snippet: Briefly, genes encoding RBD (residue 319-541) of the SARS-CoV-2 spike protein were amplified using RBD primers and codon-optimized SARS-CoV-2 Spike ORF mammalian expression plasmid (VG40589; Sino Biological, CN) as the template.

    Techniques: Mouse Assay

    W4P-RBD elicits RBD-specific antibody responses in serum and induces potent neutralizing activity against pseudotyped SARS-CoV-2. C57BL/6 mice were immunized with W-RBD, W4P-RBD (50 μ g /mouse), or empty pcDNA3.3 (Mock) three times at 1-week intervals. (A,C) Antibody responses in serum specific to SARS-CoV-2 RBD proteins were detected by ELISA. (B) Serum at 5 weeks after the last immunization was assessed using different dilution factors for IgG against the SARS-CoV-2 RBD protein using ELISA. (D) The 50% neutralizing antibody titer (NT 50 ) was calculated using the SARS-CoV-2 pseudovirus neutralization assay in Calu-3 cells. (E) Correlation between SARS-CoV-2 RBD-specific IgG and pseudotyped SARS-CoV-2 neutralization titers for immunized mice. Significance differences (* P

    Journal: Frontiers in Immunology

    Article Title: A Novel DNA Vaccine Against SARS-CoV-2 Encoding a Chimeric Protein of Its Receptor-Binding Domain (RBD) Fused to the Amino-Terminal Region of Hepatitis B Virus preS1 With a W4P Mutation

    doi: 10.3389/fimmu.2021.637654

    Figure Lengend Snippet: W4P-RBD elicits RBD-specific antibody responses in serum and induces potent neutralizing activity against pseudotyped SARS-CoV-2. C57BL/6 mice were immunized with W-RBD, W4P-RBD (50 μ g /mouse), or empty pcDNA3.3 (Mock) three times at 1-week intervals. (A,C) Antibody responses in serum specific to SARS-CoV-2 RBD proteins were detected by ELISA. (B) Serum at 5 weeks after the last immunization was assessed using different dilution factors for IgG against the SARS-CoV-2 RBD protein using ELISA. (D) The 50% neutralizing antibody titer (NT 50 ) was calculated using the SARS-CoV-2 pseudovirus neutralization assay in Calu-3 cells. (E) Correlation between SARS-CoV-2 RBD-specific IgG and pseudotyped SARS-CoV-2 neutralization titers for immunized mice. Significance differences (* P

    Article Snippet: Briefly, genes encoding RBD (residue 319-541) of the SARS-CoV-2 spike protein were amplified using RBD primers and codon-optimized SARS-CoV-2 Spike ORF mammalian expression plasmid (VG40589; Sino Biological, CN) as the template.

    Techniques: Activity Assay, Mouse Assay, Enzyme-linked Immunosorbent Assay, Neutralization

    W4P-RBD exerts potent neutralizing activity against live SARS-CoV-2. C57BL/6 mice were immunized with W-RBD, W4P-RBD (50 μ g /mouse), or empty pcDNA3.3 (Mock) three times at 1-week intervals. Serum from the immunized mice was diluted and incubated with live SARS-CoV-2 for neutralization assays. (A) A 50% plaque reduction neutralizing antibody (PRNT 50 ) titer against live SARS-CoV-2 was calculated against SARS-CoV-2 infection in Vero E6 cells. (B) Reduction in plaque formation in Vero E6 cells infected with SARS-CoV-2. (C) Correlation between SARS-CoV-2 RBD-specific IgG and SARS-CoV-2 neutralization titers in immunized mice. Significance differences (** P

    Journal: Frontiers in Immunology

    Article Title: A Novel DNA Vaccine Against SARS-CoV-2 Encoding a Chimeric Protein of Its Receptor-Binding Domain (RBD) Fused to the Amino-Terminal Region of Hepatitis B Virus preS1 With a W4P Mutation

    doi: 10.3389/fimmu.2021.637654

    Figure Lengend Snippet: W4P-RBD exerts potent neutralizing activity against live SARS-CoV-2. C57BL/6 mice were immunized with W-RBD, W4P-RBD (50 μ g /mouse), or empty pcDNA3.3 (Mock) three times at 1-week intervals. Serum from the immunized mice was diluted and incubated with live SARS-CoV-2 for neutralization assays. (A) A 50% plaque reduction neutralizing antibody (PRNT 50 ) titer against live SARS-CoV-2 was calculated against SARS-CoV-2 infection in Vero E6 cells. (B) Reduction in plaque formation in Vero E6 cells infected with SARS-CoV-2. (C) Correlation between SARS-CoV-2 RBD-specific IgG and SARS-CoV-2 neutralization titers in immunized mice. Significance differences (** P

    Article Snippet: Briefly, genes encoding RBD (residue 319-541) of the SARS-CoV-2 spike protein were amplified using RBD primers and codon-optimized SARS-CoV-2 Spike ORF mammalian expression plasmid (VG40589; Sino Biological, CN) as the template.

    Techniques: Activity Assay, Mouse Assay, Incubation, Neutralization, Plaque Reduction Neutralization Test, Infection

    The W4P-RBD vaccine exerts potent neutralizing activity against live SARS-CoV-2 and inhibits viral infection and replication of SARS-CoV-2. C57BL/6 mice were immunized with W-RBD, W4P-RBD (50 μ g /mouse), or empty pcDNA3.3 (Mock) three times at 1-week intervals. Serum from the immunized mice was diluted and incubated with live SARS-CoV-2 for neutralization assays. (A) The neutralization efficacy of serum from immunized mice against live SARS-CoV-2 RNA copy number in Vero E6 cells was determined by qRT-PCR. (B) The neutralization efficacy of the diluted serum against live SARS-CoV-2 in Vero E6 cells was determined by Western blotting. (C) Pooled serum (diluted 1:500) from each mouse group was tested in the neutralization assay against live SARS-CoV-2. Representative images (40-fold magnification) show live SARS-CoV-2-infected cells after neutralization in each group. Significance differences (* P

    Journal: Frontiers in Immunology

    Article Title: A Novel DNA Vaccine Against SARS-CoV-2 Encoding a Chimeric Protein of Its Receptor-Binding Domain (RBD) Fused to the Amino-Terminal Region of Hepatitis B Virus preS1 With a W4P Mutation

    doi: 10.3389/fimmu.2021.637654

    Figure Lengend Snippet: The W4P-RBD vaccine exerts potent neutralizing activity against live SARS-CoV-2 and inhibits viral infection and replication of SARS-CoV-2. C57BL/6 mice were immunized with W-RBD, W4P-RBD (50 μ g /mouse), or empty pcDNA3.3 (Mock) three times at 1-week intervals. Serum from the immunized mice was diluted and incubated with live SARS-CoV-2 for neutralization assays. (A) The neutralization efficacy of serum from immunized mice against live SARS-CoV-2 RNA copy number in Vero E6 cells was determined by qRT-PCR. (B) The neutralization efficacy of the diluted serum against live SARS-CoV-2 in Vero E6 cells was determined by Western blotting. (C) Pooled serum (diluted 1:500) from each mouse group was tested in the neutralization assay against live SARS-CoV-2. Representative images (40-fold magnification) show live SARS-CoV-2-infected cells after neutralization in each group. Significance differences (* P

    Article Snippet: Briefly, genes encoding RBD (residue 319-541) of the SARS-CoV-2 spike protein were amplified using RBD primers and codon-optimized SARS-CoV-2 Spike ORF mammalian expression plasmid (VG40589; Sino Biological, CN) as the template.

    Techniques: Activity Assay, Infection, Mouse Assay, Incubation, Neutralization, Quantitative RT-PCR, Western Blot

    W4P-RBD induces proinflammatory cytokine production in S1-stimulated splenocytes. Splenocytes from immunized mice were stimulated with SARS-CoV-2 S1 protein (5 μ g / ml ) for 5 days. The cytokine production of (A) TNFα, (B) IFNγ, (C) IL-12p40, (D) IFNβ, (E) IL-6, and (F) IL-2 from splenocytes was detected by ELISA. Significance differences (* P

    Journal: Frontiers in Immunology

    Article Title: A Novel DNA Vaccine Against SARS-CoV-2 Encoding a Chimeric Protein of Its Receptor-Binding Domain (RBD) Fused to the Amino-Terminal Region of Hepatitis B Virus preS1 With a W4P Mutation

    doi: 10.3389/fimmu.2021.637654

    Figure Lengend Snippet: W4P-RBD induces proinflammatory cytokine production in S1-stimulated splenocytes. Splenocytes from immunized mice were stimulated with SARS-CoV-2 S1 protein (5 μ g / ml ) for 5 days. The cytokine production of (A) TNFα, (B) IFNγ, (C) IL-12p40, (D) IFNβ, (E) IL-6, and (F) IL-2 from splenocytes was detected by ELISA. Significance differences (* P

    Article Snippet: Briefly, genes encoding RBD (residue 319-541) of the SARS-CoV-2 spike protein were amplified using RBD primers and codon-optimized SARS-CoV-2 Spike ORF mammalian expression plasmid (VG40589; Sino Biological, CN) as the template.

    Techniques: Mouse Assay, Enzyme-linked Immunosorbent Assay

    Construction of the HBV W4P preS1-fused pcDNA3.3-RBD plasmid (W4P-RBD) as a candidate for SARS-CoV-2. (A) Design of pcDNA3.3-RBD and pcDNA3.3-W4P-RBD. The W4P region comprises 33 bp from the first site of the preS1 region of the HBV genome and encodes 11 amino acids. (B) The protein expression of SARS-CoV-2 RBD and W4P-conjugated RBD was detected by the Western blot assay. pcDNA3.3-RBD, pcDNA3.3-W4P-RBD, and empty pcDNA3.3 were transfected into Vero E6, Huh7, and 293T cells, and cell lysates were collected 48 h post transfection to detect protein expression. (C) The mRNA expression levels of IL-6 and in pcDNA3.3-transfected cells were detected by qRT-PCR. Significance differences (* P

    Journal: Frontiers in Immunology

    Article Title: A Novel DNA Vaccine Against SARS-CoV-2 Encoding a Chimeric Protein of Its Receptor-Binding Domain (RBD) Fused to the Amino-Terminal Region of Hepatitis B Virus preS1 With a W4P Mutation

    doi: 10.3389/fimmu.2021.637654

    Figure Lengend Snippet: Construction of the HBV W4P preS1-fused pcDNA3.3-RBD plasmid (W4P-RBD) as a candidate for SARS-CoV-2. (A) Design of pcDNA3.3-RBD and pcDNA3.3-W4P-RBD. The W4P region comprises 33 bp from the first site of the preS1 region of the HBV genome and encodes 11 amino acids. (B) The protein expression of SARS-CoV-2 RBD and W4P-conjugated RBD was detected by the Western blot assay. pcDNA3.3-RBD, pcDNA3.3-W4P-RBD, and empty pcDNA3.3 were transfected into Vero E6, Huh7, and 293T cells, and cell lysates were collected 48 h post transfection to detect protein expression. (C) The mRNA expression levels of IL-6 and in pcDNA3.3-transfected cells were detected by qRT-PCR. Significance differences (* P

    Article Snippet: Briefly, genes encoding RBD (residue 319-541) of the SARS-CoV-2 spike protein were amplified using RBD primers and codon-optimized SARS-CoV-2 Spike ORF mammalian expression plasmid (VG40589; Sino Biological, CN) as the template.

    Techniques: Plasmid Preparation, Expressing, Western Blot, Transfection, Quantitative RT-PCR

    Surface antigenic similarity of rVSV-ΔG-spike and SARS-CoV-2: (A) Immunofluorescent images of Vero E6 cells infected with either WT-VSV (left panel), rVSV-ΔG-spike (middle panel), or SARS-CoV-2, stained with serum from COVID-19 human convalescent serum (right panel). (B) Correlation analysis of neutralization of rVSV-ΔG-spike and SARS-CoV-2 by a panel of sera from COVID-19 convalescent patients. For each sera (n=12), NT 50 values were determined for neutralization of rVSV-ΔG-spike, or SARS-CoV-2. The NT 50 values were plotted to determine the correlation between th neutralization assays. R 2 =0.911.

    Journal: bioRxiv

    Article Title: A single dose of recombinant VSV-ΔG-spike vaccine provides protection against SARS-CoV-2 challenge

    doi: 10.1101/2020.06.18.160655

    Figure Lengend Snippet: Surface antigenic similarity of rVSV-ΔG-spike and SARS-CoV-2: (A) Immunofluorescent images of Vero E6 cells infected with either WT-VSV (left panel), rVSV-ΔG-spike (middle panel), or SARS-CoV-2, stained with serum from COVID-19 human convalescent serum (right panel). (B) Correlation analysis of neutralization of rVSV-ΔG-spike and SARS-CoV-2 by a panel of sera from COVID-19 convalescent patients. For each sera (n=12), NT 50 values were determined for neutralization of rVSV-ΔG-spike, or SARS-CoV-2. The NT 50 values were plotted to determine the correlation between th neutralization assays. R 2 =0.911.

    Article Snippet: Plasmid construction The pVSV-spike expression plasmid was constructed by PCR amplification of the full length human codon optimized spike gene from pCMV3-SARS-Cov-2 spike expression plasmid (Sino Biological, Cat #VG40588-UT) using the following primers: Forward – atcgatctgtttacgcgtcactATGTTTGTGTTCCTGGTGCTGC; Reverse – atgaagaatctggctagcaggatttgagTCAGGTGTAGTGCAGTTTCACTCC.

    Techniques: Infection, Staining, Neutralization