sars cov 2 2019 ncov spike antibody rabbit pab  (Sino Biological)


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    Name:
    SARS CoV 2 2019 nCoV Spike Antibody Rabbit PAb
    Description:
    Produced in rabbits immunized with purified recombinant SARS CoV 2 2019 nCoV Spike Protein S1 S2 ECD Catalog 40589 V08B1 YP 009724390 1 Val16 Pro1213 The specific IgG was purified by SARS CoV 2 2019 nCoV Spike affinity chromatography
    Catalog Number:
    40589-T62
    Price:
    None
    Category:
    Primary Antibody
    Reactivity:
    2019 nCoV
    Applications:
    WB,ELISA
    Immunogen:
    Recombinant SARS-CoV-2 / 2019-nCoV Spike Protein (Catalog#40589-V08B1)
    Product Aliases:
    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-NCP-CoV RBD Antibody, Anti-NCP-CoV s1 Antibody, Anti-NCP-CoV s2 Antibody, Anti-NCP-CoV Spike Antibody, Anti-novel coronavirus RBD Antibody, Anti-novel coronavirus s1 Antibody, Anti-novel coronavirus s2 Antibody, Anti-novel coronavirus spike Antibody, Anti-RBD Antibody, Anti-S1 Antibody, Anti-S2 Antibody, Anti-Spike RBD Antibody
    Antibody Type:
    PAb
    Host:
    Rabbit
    Isotype:
    Rabbit IgG
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    Structured Review

    Sino Biological sars cov 2 2019 ncov spike antibody rabbit pab
    Inhibitory activity of HP-OVA against <t>SARS-CoV-2</t> S-mediated cell-cell fusion. Images were captured at 12 h after treatment with HP-OVA or OVA on SARS-CoV-2 S protein-mediated cell-cell fusion. The syncytia of Vero E6 cells (A) or Huh 7 cells (B) and HEK293T cells with SARS-CoV-2 overexpression are marked in the pictures. Representative results from three fields were selected randomly from each sample with scale bars of 50 μm (C, D) The number of syncytia was counted under an inverted fluorescence microscope, and the percentage of inhibition was calculated as described in the Methods. Data are presented as the mean ± SD of triplicate samples from a representative experiment (* p
    Produced in rabbits immunized with purified recombinant SARS CoV 2 2019 nCoV Spike Protein S1 S2 ECD Catalog 40589 V08B1 YP 009724390 1 Val16 Pro1213 The specific IgG was purified by SARS CoV 2 2019 nCoV Spike affinity chromatography
    https://www.bioz.com/result/sars cov 2 2019 ncov spike antibody rabbit pab/product/Sino Biological
    Average 96 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    sars cov 2 2019 ncov spike antibody rabbit pab - by Bioz Stars, 2021-07
    96/100 stars

    Images

    1) Product Images from "3-Hydroxyphthalic Anhydride-Modified Chicken Ovalbumin as a Potential Candidate Inhibits SARS-CoV-2 Infection by Disrupting the Interaction of Spike Protein With Host ACE2 Receptor"

    Article Title: 3-Hydroxyphthalic Anhydride-Modified Chicken Ovalbumin as a Potential Candidate Inhibits SARS-CoV-2 Infection by Disrupting the Interaction of Spike Protein With Host ACE2 Receptor

    Journal: Frontiers in Pharmacology

    doi: 10.3389/fphar.2020.603830

    Inhibitory activity of HP-OVA against SARS-CoV-2 S-mediated cell-cell fusion. Images were captured at 12 h after treatment with HP-OVA or OVA on SARS-CoV-2 S protein-mediated cell-cell fusion. The syncytia of Vero E6 cells (A) or Huh 7 cells (B) and HEK293T cells with SARS-CoV-2 overexpression are marked in the pictures. Representative results from three fields were selected randomly from each sample with scale bars of 50 μm (C, D) The number of syncytia was counted under an inverted fluorescence microscope, and the percentage of inhibition was calculated as described in the Methods. Data are presented as the mean ± SD of triplicate samples from a representative experiment (* p
    Figure Legend Snippet: Inhibitory activity of HP-OVA against SARS-CoV-2 S-mediated cell-cell fusion. Images were captured at 12 h after treatment with HP-OVA or OVA on SARS-CoV-2 S protein-mediated cell-cell fusion. The syncytia of Vero E6 cells (A) or Huh 7 cells (B) and HEK293T cells with SARS-CoV-2 overexpression are marked in the pictures. Representative results from three fields were selected randomly from each sample with scale bars of 50 μm (C, D) The number of syncytia was counted under an inverted fluorescence microscope, and the percentage of inhibition was calculated as described in the Methods. Data are presented as the mean ± SD of triplicate samples from a representative experiment (* p

    Techniques Used: Activity Assay, Over Expression, Fluorescence, Microscopy, Inhibition

    Inhibition of HP-OVA on the infection with SARS-CoV-2 PsV and SARS-CoV PsV. Antiviral activity of HP-OVA against SARS-CoV-2 S PsV infection in 293T/ACE2 (A) or Vero E6 (B) target cells. Inhibition of single-round infection of SARS-CoV S PsV in 293T/ACE2 (C) and Vero E6 (D) cells. Data are presented as the mean ± SD of triplicate samples from a representative experiment (* p
    Figure Legend Snippet: Inhibition of HP-OVA on the infection with SARS-CoV-2 PsV and SARS-CoV PsV. Antiviral activity of HP-OVA against SARS-CoV-2 S PsV infection in 293T/ACE2 (A) or Vero E6 (B) target cells. Inhibition of single-round infection of SARS-CoV S PsV in 293T/ACE2 (C) and Vero E6 (D) cells. Data are presented as the mean ± SD of triplicate samples from a representative experiment (* p

    Techniques Used: Inhibition, Infection, Activity Assay

    HP-OVA binding to both SARS-CoV-2 S and ACE2 protein. Analysis of the expression of SARS-CoV-2 S (A) and ACE2 (B) in HEK-293T cells by western blot. The binding of HP-OVA to cells expressing SARS-CoV-2 S (C) or ACE2 (D) was assessed by flow cytometry. A representative flow histogram and quantification of the binding of HP-OVA to cells expressing SARS-CoV-2 S (E) or ACE2 (F) were shown. Data are presented as the mean ± SD (* p
    Figure Legend Snippet: HP-OVA binding to both SARS-CoV-2 S and ACE2 protein. Analysis of the expression of SARS-CoV-2 S (A) and ACE2 (B) in HEK-293T cells by western blot. The binding of HP-OVA to cells expressing SARS-CoV-2 S (C) or ACE2 (D) was assessed by flow cytometry. A representative flow histogram and quantification of the binding of HP-OVA to cells expressing SARS-CoV-2 S (E) or ACE2 (F) were shown. Data are presented as the mean ± SD (* p

    Techniques Used: Binding Assay, Expressing, Western Blot, Flow Cytometry

    The interaction of HP-OVA with SARS-CoV-2 S and ACE2. The binding of OVA to SARS-CoV-2 spike (RBD), S2 and BSA protein was assessed by ELISA (A) . The binding of HP-OVA to SARS-CoV-2 spike (RBD), S2 and a negative control BSA protein was assessed by ELISA (B) . The binding ability of HP-OVA to ACE2 protein was assessed by ELISA (C) . Inhibition of the interaction between spike (RBD) and ACE2 proteins by HP-OVA, as determined by a competitive inhibition ELISA (D) . Data are presented as the mean ± SD of triplicate samples from a representative experiment (* p
    Figure Legend Snippet: The interaction of HP-OVA with SARS-CoV-2 S and ACE2. The binding of OVA to SARS-CoV-2 spike (RBD), S2 and BSA protein was assessed by ELISA (A) . The binding of HP-OVA to SARS-CoV-2 spike (RBD), S2 and a negative control BSA protein was assessed by ELISA (B) . The binding ability of HP-OVA to ACE2 protein was assessed by ELISA (C) . Inhibition of the interaction between spike (RBD) and ACE2 proteins by HP-OVA, as determined by a competitive inhibition ELISA (D) . Data are presented as the mean ± SD of triplicate samples from a representative experiment (* p

    Techniques Used: Binding Assay, Enzyme-linked Immunosorbent Assay, Negative Control, Inhibition

    Schematic representation of the molecular mechanisms of HP-OVA against SARS-CoV-2 infection. HP-OVA binds to both the S protein of SARS-CoV-2 and host angiotensin-converting enzyme 2 (ACE2), the functional receptor of SARS-CoV-2, and disrupts the S protein-ACE2 interaction, thereby exhibiting inhibitory activity against SARS-CoV-2 infection.
    Figure Legend Snippet: Schematic representation of the molecular mechanisms of HP-OVA against SARS-CoV-2 infection. HP-OVA binds to both the S protein of SARS-CoV-2 and host angiotensin-converting enzyme 2 (ACE2), the functional receptor of SARS-CoV-2, and disrupts the S protein-ACE2 interaction, thereby exhibiting inhibitory activity against SARS-CoV-2 infection.

    Techniques Used: Infection, Functional Assay, Activity Assay

    Related Articles

    Enzyme-linked Immunosorbent Assay:

    Article Title: Rapid and sensitive detection of SARS-CoV-2 antibodies by biolayer interferometry
    Article Snippet: Finally, we believe that BLI-ISA can be developed as a novel diagnostic platform to evaluate antibodies and other biomolecules in clinical specimens, for example to evaluate plasma antibody levels to inform patients on vaccinations, or to quickly identify and prioritize donors for convalescent plasma therapy donation , . .. Reagents and suppliesPhosphate buffered saline (PBS) tablets (Sigma P4417), Tween-20 (Fisher BP337), dry milk powder (RPI 50488786), ELISA plates (Corning 3590), Goat anti-Human IgG Fc HRP (Thermo Fisher A18817), OPD tablets (Pierce PI34006), bovine serum albumin (BSA) (Fisher BP1600), ChonBlock (Chondrex 9068), Biotinylated SARS-CoV-2 protein RBD His AviTag (Acro Biosystems SPD-C82E9), 4 nm Colloidal Gold-AffiPure Goat Anti-Human IgG Fcg fragment specific (Jackson ImmunoResearch 109-185-098) rehydrated in 1 mL deionized water per the manufacturer’s instructions, 4 nm Colloidal Gold-AffiPure Goat Anti-Human Serum IgA alpha chain specific (Jackson ImmunoResearch 109-185-011) rehydrated in 1 mL deionized water per the manufacturer’s instructions, Human coronavirus spike glycoprotein Antibody, Rabbit PAb, Antigen Affinity Purified (Sino Biological 40021-T60), SARS-CoV-2 (2019-nCoV) spike Antibody, Rabbit PAb, Antigen Affinity Purified (Sino Biological 40589-T62), SARS-CoV-2 (2019-nCoV) spike Antibody, Rabbit MAb (40150-R007), Anti-SARS-CoV S Therapeutic Antibody (CR3022) (Creative Biolabs MRO-1214LC), Octet Anti-Penta-His (HIS1K) sensor tips (Sartorius ForteBio 18-5120), Octet Streptavidin (SA) sensor tips (Sartorius ForteBio 18-5019), tilted bottom (TW384) microplates (Sartorius ForteBio 18-5080), electroporation cuvettes (MaxCyte SOC4), suspension adapted CHO-S cells (Thermo Fisher R80007). .. CD-CHO medium (Thermo Fisher 10743029), CD OptiCHO medium (Thermo Fisher 12681011), HisTrap FF (GE Healthcare 17-5286-01), StrepTrap HP (GE Healthcare 28-9075-47), Superdex 200 Increase GL (GE Healthcare 28-9909-44).

    Article Title: 3-Hydroxyphthalic Anhydride-Modified Chicken Ovalbumin as a Potential Candidate Inhibits SARS-CoV-2 Infection by Disrupting the Interaction of Spike Protein With Host ACE2 Receptor
    Article Snippet: Unmodified OVA was used as a negative control. .. Enzyme-Linked Immunosorbent Assay (ELISA)ELISA was performed to identify the interaction of HP-OVA and the SARS-CoV-2 S protein (RBD) or ACE2 protein. .. Briefly, wells of 96-well polystyrene microplates were coated with 1 μg/mL S protein (RBD) (Sino Biological, China) or ACE2 protein (Invitrogen, Carlsbad, CA) in 0.01 M Tris buffer (pH 8.8) at 4°C overnight.

    Affinity Purification:

    Article Title: Rapid and sensitive detection of SARS-CoV-2 antibodies by biolayer interferometry
    Article Snippet: Finally, we believe that BLI-ISA can be developed as a novel diagnostic platform to evaluate antibodies and other biomolecules in clinical specimens, for example to evaluate plasma antibody levels to inform patients on vaccinations, or to quickly identify and prioritize donors for convalescent plasma therapy donation , . .. Reagents and suppliesPhosphate buffered saline (PBS) tablets (Sigma P4417), Tween-20 (Fisher BP337), dry milk powder (RPI 50488786), ELISA plates (Corning 3590), Goat anti-Human IgG Fc HRP (Thermo Fisher A18817), OPD tablets (Pierce PI34006), bovine serum albumin (BSA) (Fisher BP1600), ChonBlock (Chondrex 9068), Biotinylated SARS-CoV-2 protein RBD His AviTag (Acro Biosystems SPD-C82E9), 4 nm Colloidal Gold-AffiPure Goat Anti-Human IgG Fcg fragment specific (Jackson ImmunoResearch 109-185-098) rehydrated in 1 mL deionized water per the manufacturer’s instructions, 4 nm Colloidal Gold-AffiPure Goat Anti-Human Serum IgA alpha chain specific (Jackson ImmunoResearch 109-185-011) rehydrated in 1 mL deionized water per the manufacturer’s instructions, Human coronavirus spike glycoprotein Antibody, Rabbit PAb, Antigen Affinity Purified (Sino Biological 40021-T60), SARS-CoV-2 (2019-nCoV) spike Antibody, Rabbit PAb, Antigen Affinity Purified (Sino Biological 40589-T62), SARS-CoV-2 (2019-nCoV) spike Antibody, Rabbit MAb (40150-R007), Anti-SARS-CoV S Therapeutic Antibody (CR3022) (Creative Biolabs MRO-1214LC), Octet Anti-Penta-His (HIS1K) sensor tips (Sartorius ForteBio 18-5120), Octet Streptavidin (SA) sensor tips (Sartorius ForteBio 18-5019), tilted bottom (TW384) microplates (Sartorius ForteBio 18-5080), electroporation cuvettes (MaxCyte SOC4), suspension adapted CHO-S cells (Thermo Fisher R80007). .. CD-CHO medium (Thermo Fisher 10743029), CD OptiCHO medium (Thermo Fisher 12681011), HisTrap FF (GE Healthcare 17-5286-01), StrepTrap HP (GE Healthcare 28-9075-47), Superdex 200 Increase GL (GE Healthcare 28-9909-44).

    Electroporation:

    Article Title: Rapid and sensitive detection of SARS-CoV-2 antibodies by biolayer interferometry
    Article Snippet: Finally, we believe that BLI-ISA can be developed as a novel diagnostic platform to evaluate antibodies and other biomolecules in clinical specimens, for example to evaluate plasma antibody levels to inform patients on vaccinations, or to quickly identify and prioritize donors for convalescent plasma therapy donation , . .. Reagents and suppliesPhosphate buffered saline (PBS) tablets (Sigma P4417), Tween-20 (Fisher BP337), dry milk powder (RPI 50488786), ELISA plates (Corning 3590), Goat anti-Human IgG Fc HRP (Thermo Fisher A18817), OPD tablets (Pierce PI34006), bovine serum albumin (BSA) (Fisher BP1600), ChonBlock (Chondrex 9068), Biotinylated SARS-CoV-2 protein RBD His AviTag (Acro Biosystems SPD-C82E9), 4 nm Colloidal Gold-AffiPure Goat Anti-Human IgG Fcg fragment specific (Jackson ImmunoResearch 109-185-098) rehydrated in 1 mL deionized water per the manufacturer’s instructions, 4 nm Colloidal Gold-AffiPure Goat Anti-Human Serum IgA alpha chain specific (Jackson ImmunoResearch 109-185-011) rehydrated in 1 mL deionized water per the manufacturer’s instructions, Human coronavirus spike glycoprotein Antibody, Rabbit PAb, Antigen Affinity Purified (Sino Biological 40021-T60), SARS-CoV-2 (2019-nCoV) spike Antibody, Rabbit PAb, Antigen Affinity Purified (Sino Biological 40589-T62), SARS-CoV-2 (2019-nCoV) spike Antibody, Rabbit MAb (40150-R007), Anti-SARS-CoV S Therapeutic Antibody (CR3022) (Creative Biolabs MRO-1214LC), Octet Anti-Penta-His (HIS1K) sensor tips (Sartorius ForteBio 18-5120), Octet Streptavidin (SA) sensor tips (Sartorius ForteBio 18-5019), tilted bottom (TW384) microplates (Sartorius ForteBio 18-5080), electroporation cuvettes (MaxCyte SOC4), suspension adapted CHO-S cells (Thermo Fisher R80007). .. CD-CHO medium (Thermo Fisher 10743029), CD OptiCHO medium (Thermo Fisher 12681011), HisTrap FF (GE Healthcare 17-5286-01), StrepTrap HP (GE Healthcare 28-9075-47), Superdex 200 Increase GL (GE Healthcare 28-9909-44).

    Infection:

    Article Title: Large scale discovery of coronavirus-host factor protein interaction motifs reveals SARS-CoV-2 specific mechanisms and vulnerabilities
    Article Snippet: .. VeroE6 cells were seeded in 8-well chamber slides (Sarstedts) and infected with SARS CoV-2 for 6 h. Cells were fixed with 4% formaldehyde, quenched with 10 mM glycine, and permeabilized with PBS and 0.5% Triton X-100. .. Thereafter, cells are incubated with primary antibodies against SARS-CoV-2 nucleocapsid ((1:500) Sino Biological Inc., 40143-R001) and G3BP1 ((1:500) Abcam, ab56574) followed by incubation with conjugated secondary antibodies anti-rabbit Alexa555 and anti-mouse Alexa488 (1:500, Thermo Fisher Scientific).

    Article Title: Prime-boost vaccination of mice and Rhesus macaques with two novel adenovirus vectored COVID-19 vaccine candidates
    Article Snippet: Western blottingHEK-293A cells were infected with Sad23L-nCoV-S and Ad49L-nCoV-S strains, respectively, and Sad23L-GFP and Ad49L-GFP vectorial viruses were used as mock control. .. The expression of SARS-CoV-2 S protein was analyzed by Western blotting with rabbit polyclonal antibody to SARS-CoV-2 RBD (Sino Biological, China) and heat-inactivated human serum samples from Chinese COVID-19 infected patients. ..

    Expressing:

    Article Title: Prime-boost vaccination of mice and rhesus macaques with two novel adenovirus vectored COVID-19 vaccine candidates
    Article Snippet: Western blottingHEK-293A cells were infected with Sad23L-nCoV-S and Ad49L-nCoV-S strains, respectively, and Sad23L-GFP and Ad49L-GFP vectorial viruses were used as mock control. .. The expression of SARS-CoV-2 S protein was analysed by Western blotting with rabbit polyclonal antibody to SARS-CoV-2 RBD (Sino Biological, 40592-T62, China). ..

    Article Title: Prime-boost vaccination of mice and Rhesus macaques with two novel adenovirus vectored COVID-19 vaccine candidates
    Article Snippet: Western blottingHEK-293A cells were infected with Sad23L-nCoV-S and Ad49L-nCoV-S strains, respectively, and Sad23L-GFP and Ad49L-GFP vectorial viruses were used as mock control. .. The expression of SARS-CoV-2 S protein was analyzed by Western blotting with rabbit polyclonal antibody to SARS-CoV-2 RBD (Sino Biological, China) and heat-inactivated human serum samples from Chinese COVID-19 infected patients. ..

    Western Blot:

    Article Title: Prime-boost vaccination of mice and rhesus macaques with two novel adenovirus vectored COVID-19 vaccine candidates
    Article Snippet: Western blottingHEK-293A cells were infected with Sad23L-nCoV-S and Ad49L-nCoV-S strains, respectively, and Sad23L-GFP and Ad49L-GFP vectorial viruses were used as mock control. .. The expression of SARS-CoV-2 S protein was analysed by Western blotting with rabbit polyclonal antibody to SARS-CoV-2 RBD (Sino Biological, 40592-T62, China). ..

    Article Title: Prime-boost vaccination of mice and Rhesus macaques with two novel adenovirus vectored COVID-19 vaccine candidates
    Article Snippet: Western blottingHEK-293A cells were infected with Sad23L-nCoV-S and Ad49L-nCoV-S strains, respectively, and Sad23L-GFP and Ad49L-GFP vectorial viruses were used as mock control. .. The expression of SARS-CoV-2 S protein was analyzed by Western blotting with rabbit polyclonal antibody to SARS-CoV-2 RBD (Sino Biological, China) and heat-inactivated human serum samples from Chinese COVID-19 infected patients. ..

    Article Title: Structural basis for enhanced infectivity and immune evasion of SARS-CoV-2 variants
    Article Snippet: The dimeric ACE2 protein was purified by GammaBind Plus Sepharose beads (GE Healthcare), followed gel filtration chromatography on a Superdex 200 Increase 10/300 GL column. .. Western blot Western blot was performed using an anti-SARS-COV-2 S antibody following a protocol described previously . .. Briefly, full-length S protein samples were prepared from cell pellets and resolved in 4-15% Mini-Protean TGX gel (Bio-Rad, Hercules, CA) and transferred onto PVDF membranes.

    Neutralization:

    Article Title: Characterization of spike glycoprotein of SARS-CoV-2 on virus entry and its immune cross-reactivity with SARS-CoV
    Article Snippet: Pseudoviral transduction was measured according to luciferase activities. .. Pseudovirus neutralization assaySARS-CoV S, SARS-CoV-2 S, and VSV-G pseudovirions were pre-incubated with serially diluted either polyclonal rabbit anti-SARS S1 antibodies T62 or patient sera for 1 h on ice, then virus-antibody mixture was added onto 293/hACE2 cells in a 96-well plate. .. Cells were lysed 40 h later and pseudovirus transduction was measured as previously described.

    Staining:

    Article Title: A human monoclonal antibody blocking SARS-CoV-2 infection
    Article Snippet: The mixture was then added to VeroE6 cells and incubated for 1 hour, after which the cells were washed and further incubated in medium for 8 hours. .. The cells were then fixed and stained using a rabbit anti-SARS-CoV serum (Sino Biological) and a secondary peroxidase-labeled goat anti-rabbit IgG (Dako). .. The signal was developed using a precipitate forming TMB substrate (True Blue, KPL) and the number of infected cells per well were counted using the ImmunoSpot Image analyzer (CTL Europe GmbH).

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    Sino Biological sars cov 2 2019 ncov spike rbd antibody rabbit pab
    ELISA ( x -axis) vs. LFRET ( y -axis) results by disease severity. ( a ) Anti-NP IgA ELISA vs. anti-NP LFRET (N = 81, R = 0.25). ( b ) anti-NP IgG ELISA vs. anti-NP LFRET (N = 129, R = 0.62). ( c ) anti-NP IgM ELISA vs. anti-NP LFRET (N = 81, R = 0.13). ( d ) anti-SP IgA ELISA vs. anti-SP LFRET (N = 129, R = 0.53). ( e ) anti-SP IgG ELISA vs. anti-SP LFRET (N = 129, R = 0.62). ( f ) anti-SP IgM ELISA vs. anti-SP LFRET (N = 81, R = 0.56). Color of the dot indicates <t>SARS-CoV-2</t> PCR result and disease severity: cyan = PCR negative; yellow = non-hospitalized, PCR-positive; red = non-ICU hospitalized, PCR positive; black = hospitalized in ICU, PCR positive. Horizontal and vertical black lines indicate LFRET and ELISA cutoffs. On the x -axis, ELISA absorbance on a logarithmic scale and on the y -axis, LFRET signal on a logarithmic scale. SP = spike glycoprotein. NP = nucleoprotein. LFRET = protein L–based time-resolved Förster resonance energy transfer immunoassay. ELISA = enzyme immunoassay. R = Pearson’s correlation coefficient.
    Sars Cov 2 2019 Ncov Spike Rbd Antibody Rabbit Pab, 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/sars cov 2 2019 ncov spike rbd antibody rabbit pab/product/Sino Biological
    Average 99 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    sars cov 2 2019 ncov spike rbd antibody rabbit pab - by Bioz Stars, 2021-07
    99/100 stars
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    97
    Sino Biological sars cov 2 rbd
    ACE2 is required for the cellular uptake and targeting of <t>SARS-CoV-2</t> S protein RBD-tagged EVs. (A)Western blot analysis of human ACE2 was performed with cell lysates from various cell lines or organoid as where indicated using antibody against ACE2. (B) Schematic illustration of RBD-GFP-loaded EVs. Cultured cells were transfected with RBD-VSVG vector or empty vector (NC-vector) in combination with CD9-GFP vector. 48 h post transfection, the EV fraction that was labeled with GFP was collected(EV-RBD-GFP + /EV-NC-GFP + ). (C) Flow cytometric analysis of GFP ratio in different cell lines after incubating with 1 × 10 9 of EV-RBD-GFP+ or EV-NC-GFP+ for 6 h. Different symbols correspond to independent experiments. Data are shown as mean ± SEM of three independent experiments, each individual experiment performs triplicate. (D, E and F) Representative images of Caco-2 cells (D), iPS cells-derived cardiomyocytes (E) and intestinal organoids (F) after application of 1 × 10 9 EV-RBD-GFP + or EV-NC-GFP + for 6 h. The GFP ratio was determined by flow cytometric analysis. (G) The binding affinity for RBD-tagged EVs with ACE2 protein was quantified by ELISA. Different concentration of EVs were incubated and followed by the anti-CD9 as the detector antibody. Each concentration was repeated with six wells. Data are shown as mean ± SEM of three independent experiments. (H and I) Representative images of ACE2-A549 cells showed an earlier plasma membrane distribution of EV-RBD-GFP + or EV-NC-GFP + incubation for 1 h (H) and 6 h (I) with or without pre-treated with ACE2 antibody. Fluorescence intensity of GFP was analyzed by ImageJ software were averaged from six different fields for each. Data are shown as mean ± SEM of three independent experiments. ** p
    Sars Cov 2 Rbd, supplied by Sino Biological, used in various techniques. Bioz Stars score: 97/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/sars cov 2 rbd/product/Sino Biological
    Average 97 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    sars cov 2 rbd - by Bioz Stars, 2021-07
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    ELISA ( x -axis) vs. LFRET ( y -axis) results by disease severity. ( a ) Anti-NP IgA ELISA vs. anti-NP LFRET (N = 81, R = 0.25). ( b ) anti-NP IgG ELISA vs. anti-NP LFRET (N = 129, R = 0.62). ( c ) anti-NP IgM ELISA vs. anti-NP LFRET (N = 81, R = 0.13). ( d ) anti-SP IgA ELISA vs. anti-SP LFRET (N = 129, R = 0.53). ( e ) anti-SP IgG ELISA vs. anti-SP LFRET (N = 129, R = 0.62). ( f ) anti-SP IgM ELISA vs. anti-SP LFRET (N = 81, R = 0.56). Color of the dot indicates SARS-CoV-2 PCR result and disease severity: cyan = PCR negative; yellow = non-hospitalized, PCR-positive; red = non-ICU hospitalized, PCR positive; black = hospitalized in ICU, PCR positive. Horizontal and vertical black lines indicate LFRET and ELISA cutoffs. On the x -axis, ELISA absorbance on a logarithmic scale and on the y -axis, LFRET signal on a logarithmic scale. SP = spike glycoprotein. NP = nucleoprotein. LFRET = protein L–based time-resolved Förster resonance energy transfer immunoassay. ELISA = enzyme immunoassay. R = Pearson’s correlation coefficient.

    Journal: Viruses

    Article Title: A 10-Minute “Mix and Read” Antibody Assay for SARS-CoV-2

    doi: 10.3390/v13020143

    Figure Lengend Snippet: ELISA ( x -axis) vs. LFRET ( y -axis) results by disease severity. ( a ) Anti-NP IgA ELISA vs. anti-NP LFRET (N = 81, R = 0.25). ( b ) anti-NP IgG ELISA vs. anti-NP LFRET (N = 129, R = 0.62). ( c ) anti-NP IgM ELISA vs. anti-NP LFRET (N = 81, R = 0.13). ( d ) anti-SP IgA ELISA vs. anti-SP LFRET (N = 129, R = 0.53). ( e ) anti-SP IgG ELISA vs. anti-SP LFRET (N = 129, R = 0.62). ( f ) anti-SP IgM ELISA vs. anti-SP LFRET (N = 81, R = 0.56). Color of the dot indicates SARS-CoV-2 PCR result and disease severity: cyan = PCR negative; yellow = non-hospitalized, PCR-positive; red = non-ICU hospitalized, PCR positive; black = hospitalized in ICU, PCR positive. Horizontal and vertical black lines indicate LFRET and ELISA cutoffs. On the x -axis, ELISA absorbance on a logarithmic scale and on the y -axis, LFRET signal on a logarithmic scale. SP = spike glycoprotein. NP = nucleoprotein. LFRET = protein L–based time-resolved Förster resonance energy transfer immunoassay. ELISA = enzyme immunoassay. R = Pearson’s correlation coefficient.

    Article Snippet: At 48 h, the medium was analyzed for the presence of SARS-CoV-2 SP by dot blotting; briefly via drying 2.5 µL of the supernatant onto a nitrocellulose membrane, which then was blocked (3% skim milk in Tris-buffered saline with 0.05% Tween-20), washed, probed with rabbit anti-RBD (40592-T62, Sino Biological, Beijing, China), washed, probed with anti-rabbit IRDye800 (LI-COR Biosciences, Lincoln, NE, USA), washed, and read using Odyssey Infrared Imaging System (LI-COR Biosciences).

    Techniques: Enzyme-linked Immunosorbent Assay, Polymerase Chain Reaction, Förster Resonance Energy Transfer

    Microneutralization vs. LFRET and ELISA. Microneutralization titers are on the x -axis and LFRET signal or ELISA absorbance on the y -axis. Logarithmic scale is used on both axes. ( a ) Microneutralization titer vs. anti-SP LFRET signal (N = 107, ρ = 0.87). ( b – d ) Microneutralization titer vs. anti-SP IgG, IgA and IgM ELISA (N = 107, 107 and 67, ρ = 0.68, 0.86 and 0.81). ( e ) Microneutralization titer vs. anti-NP LFRET signal (N = 107, ρ = 0.83). ( f – h ) Microneutralization titer vs. anti-NP IgG, IgA and IgM ELISA (N = 107, 67 and 67, ρ = 0.81, 0.69 and 0.61). Color of the dots indicate SARS-CoV-2 PCR result and disease severity: cyan = PCR negative; yellow = non-hospitalized, PCR-positive; red = non-ICU hospitalized, PCR positive; black = hospitalized in ICU, PCR positive. Horizontal black lines indicate LFRET/ELISA cutoffs. SP = spike glycoprotein. NP = nucleoprotein. LFRET = protein L–based time-resolved Förster resonance energy transfer immunoassay. ELISA = enzyme immunoassay. ρ = Spearman’s rank correlation coefficient.

    Journal: Viruses

    Article Title: A 10-Minute “Mix and Read” Antibody Assay for SARS-CoV-2

    doi: 10.3390/v13020143

    Figure Lengend Snippet: Microneutralization vs. LFRET and ELISA. Microneutralization titers are on the x -axis and LFRET signal or ELISA absorbance on the y -axis. Logarithmic scale is used on both axes. ( a ) Microneutralization titer vs. anti-SP LFRET signal (N = 107, ρ = 0.87). ( b – d ) Microneutralization titer vs. anti-SP IgG, IgA and IgM ELISA (N = 107, 107 and 67, ρ = 0.68, 0.86 and 0.81). ( e ) Microneutralization titer vs. anti-NP LFRET signal (N = 107, ρ = 0.83). ( f – h ) Microneutralization titer vs. anti-NP IgG, IgA and IgM ELISA (N = 107, 67 and 67, ρ = 0.81, 0.69 and 0.61). Color of the dots indicate SARS-CoV-2 PCR result and disease severity: cyan = PCR negative; yellow = non-hospitalized, PCR-positive; red = non-ICU hospitalized, PCR positive; black = hospitalized in ICU, PCR positive. Horizontal black lines indicate LFRET/ELISA cutoffs. SP = spike glycoprotein. NP = nucleoprotein. LFRET = protein L–based time-resolved Förster resonance energy transfer immunoassay. ELISA = enzyme immunoassay. ρ = Spearman’s rank correlation coefficient.

    Article Snippet: At 48 h, the medium was analyzed for the presence of SARS-CoV-2 SP by dot blotting; briefly via drying 2.5 µL of the supernatant onto a nitrocellulose membrane, which then was blocked (3% skim milk in Tris-buffered saline with 0.05% Tween-20), washed, probed with rabbit anti-RBD (40592-T62, Sino Biological, Beijing, China), washed, probed with anti-rabbit IRDye800 (LI-COR Biosciences, Lincoln, NE, USA), washed, and read using Odyssey Infrared Imaging System (LI-COR Biosciences).

    Techniques: Enzyme-linked Immunosorbent Assay, Polymerase Chain Reaction, Förster Resonance Energy Transfer

    Simplified protocol for SARS-CoV-2 NP and SP LFRET assay. Eu-NP/-SP = Europium-labeled nucleoprotein/spike glycoprotein. AF-L = Alexa Fluor™ 647 -labeled protein L. TR-FRET = time-resolved Förster resonance energy transfer. RT = room temperature. TBS+BSA (50 mM Tris-HCl, 150 mM NaCl, pH 7.4, 0.2% BSA) was used for all dilutions. On-plate dilutions were 5 nM Eu-NP/500 nM AF-L/serum 1/25 for anti-NP and 5 nM Eu-SP/250 nM AF-L/serum 1/100 for anti-SP LFRET. For further details see the prior publication [ 5 ].

    Journal: Viruses

    Article Title: A 10-Minute “Mix and Read” Antibody Assay for SARS-CoV-2

    doi: 10.3390/v13020143

    Figure Lengend Snippet: Simplified protocol for SARS-CoV-2 NP and SP LFRET assay. Eu-NP/-SP = Europium-labeled nucleoprotein/spike glycoprotein. AF-L = Alexa Fluor™ 647 -labeled protein L. TR-FRET = time-resolved Förster resonance energy transfer. RT = room temperature. TBS+BSA (50 mM Tris-HCl, 150 mM NaCl, pH 7.4, 0.2% BSA) was used for all dilutions. On-plate dilutions were 5 nM Eu-NP/500 nM AF-L/serum 1/25 for anti-NP and 5 nM Eu-SP/250 nM AF-L/serum 1/100 for anti-SP LFRET. For further details see the prior publication [ 5 ].

    Article Snippet: At 48 h, the medium was analyzed for the presence of SARS-CoV-2 SP by dot blotting; briefly via drying 2.5 µL of the supernatant onto a nitrocellulose membrane, which then was blocked (3% skim milk in Tris-buffered saline with 0.05% Tween-20), washed, probed with rabbit anti-RBD (40592-T62, Sino Biological, Beijing, China), washed, probed with anti-rabbit IRDye800 (LI-COR Biosciences, Lincoln, NE, USA), washed, and read using Odyssey Infrared Imaging System (LI-COR Biosciences).

    Techniques: Labeling, Förster Resonance Energy Transfer

    T cell response in mice after immunization with pSARS-S and SARS2-S DNA vaccines. BALB/c (A-D) and C57BL/6 (E-H) mice (n = 4 per group) were intramuscularly immunized twice at a 3-week interval with 100 μg of vector, pSARS-S or pSARS2-S, followed by electroporation. Splenocytes were collected at week 4 after the first immunization, and the levels of secreted IFN-γ (A, E), IL-2 (B, F), IL-5 (C, G) and IL-13 (D, H) were evaluated after restimulation with recombinant SARS-CoV-2 S protein. Antibody titers are presented as the mean ± SEM. *p

    Journal: PLoS Neglected Tropical Diseases

    Article Title: DNA vaccination induced protective immunity against SARS CoV-2 infection in hamsterss

    doi: 10.1371/journal.pntd.0009374

    Figure Lengend Snippet: T cell response in mice after immunization with pSARS-S and SARS2-S DNA vaccines. BALB/c (A-D) and C57BL/6 (E-H) mice (n = 4 per group) were intramuscularly immunized twice at a 3-week interval with 100 μg of vector, pSARS-S or pSARS2-S, followed by electroporation. Splenocytes were collected at week 4 after the first immunization, and the levels of secreted IFN-γ (A, E), IL-2 (B, F), IL-5 (C, G) and IL-13 (D, H) were evaluated after restimulation with recombinant SARS-CoV-2 S protein. Antibody titers are presented as the mean ± SEM. *p

    Article Snippet: The proteins were then transferred to PVDF membranes and blotted with rabbit anti-Spike polyclonal antibody (40592-T62, Sino Biological).

    Techniques: Mouse Assay, Plasmid Preparation, Electroporation, Recombinant

    Antibody response in mice after immunization with SARS-CoV and SARS-CoV-2 S DNA vaccines. (A) BALB/c mice (n = 4 per group) were intramuscularly immunized twice at a 3-week interval with 100 μg of indicated plasmid, followed by electroporation. Serum samples were collected at the indicated time points after the first immunization. (B-D, F, G) Antibodies against the SARS-CoV-2 full-length spike protein, S2 region and RBD were evaluated by ELISA. (E, H) Vaccine-induced neutralizing antibody against SARS-CoV-2 was evaluated by neutralization assay. Antibody titers are presented as the mean ± SEM, and neutralization titers are expressed as the geometric mean with a 95% confidence interval. *p

    Journal: PLoS Neglected Tropical Diseases

    Article Title: DNA vaccination induced protective immunity against SARS CoV-2 infection in hamsterss

    doi: 10.1371/journal.pntd.0009374

    Figure Lengend Snippet: Antibody response in mice after immunization with SARS-CoV and SARS-CoV-2 S DNA vaccines. (A) BALB/c mice (n = 4 per group) were intramuscularly immunized twice at a 3-week interval with 100 μg of indicated plasmid, followed by electroporation. Serum samples were collected at the indicated time points after the first immunization. (B-D, F, G) Antibodies against the SARS-CoV-2 full-length spike protein, S2 region and RBD were evaluated by ELISA. (E, H) Vaccine-induced neutralizing antibody against SARS-CoV-2 was evaluated by neutralization assay. Antibody titers are presented as the mean ± SEM, and neutralization titers are expressed as the geometric mean with a 95% confidence interval. *p

    Article Snippet: The proteins were then transferred to PVDF membranes and blotted with rabbit anti-Spike polyclonal antibody (40592-T62, Sino Biological).

    Techniques: Mouse Assay, Plasmid Preparation, Electroporation, Enzyme-linked Immunosorbent Assay, Neutralization

    Competitive activity of immunized mouse sera against the RBD/ACE2 interaction. BALB/c mice (n = 4 per group) were intramuscularly immunized twice at a 3-week interval with 100 μg of vector, pSARS-S or pSARS2-S, followed by electroporation. Serum samples were collected at week 8 after the first immunization. Serum antibodies that compete with ACE2 for RBD binding were evaluated by competitive SARS-CoV-2 serology assay. The competitive activity of the mouse sera is expressed as the equivalent level of anti-RBD (SARS-CoV-2 spike protein) antibody (reference antibody). Antibody titers are presented as the mean ± SEM. *p

    Journal: PLoS Neglected Tropical Diseases

    Article Title: DNA vaccination induced protective immunity against SARS CoV-2 infection in hamsterss

    doi: 10.1371/journal.pntd.0009374

    Figure Lengend Snippet: Competitive activity of immunized mouse sera against the RBD/ACE2 interaction. BALB/c mice (n = 4 per group) were intramuscularly immunized twice at a 3-week interval with 100 μg of vector, pSARS-S or pSARS2-S, followed by electroporation. Serum samples were collected at week 8 after the first immunization. Serum antibodies that compete with ACE2 for RBD binding were evaluated by competitive SARS-CoV-2 serology assay. The competitive activity of the mouse sera is expressed as the equivalent level of anti-RBD (SARS-CoV-2 spike protein) antibody (reference antibody). Antibody titers are presented as the mean ± SEM. *p

    Article Snippet: The proteins were then transferred to PVDF membranes and blotted with rabbit anti-Spike polyclonal antibody (40592-T62, Sino Biological).

    Techniques: Activity Assay, Mouse Assay, Plasmid Preparation, Electroporation, Binding Assay

    Prophylactic efficacy of SARS-CoV-2 S DNA vaccine in SARS-CoV-2-infected hamsters. (A) Time course of DNA vaccination and SARS-CoV-2 challenge. Syrian hamsters were intramuscularly immunized twice at a 3-week interval with 100 μg of control, pSARS-S or pSARS2-S, followed by electroporation. Serum samples were collected by retroorbital blood sampling at weeks 4 and 6 after the first immunization. At 4 weeks after the second immunization, Syrian hamsters were intranasally challenged with 10 5 TCID 50 SARS-CoV-2. (B) Antibodies against the SARS-CoV-2 full-length spike protein were evaluated by ELISA. (C) Vaccine-induced neutralizing activity against SARS-CoV-2 was evaluated by neutralization assay. (D) Body weight change (%) of the hamsters was recorded every day after SARS-CoV-2 challenge. Virus titers (E) and viral RNA copies (F) in the lungs of SARS-CoV-2-infected hamsters at 3 days postchallenge were determined by TCID 50 assay and qRT-PCR, respectively. Antibody titers are presented as the mean ± SEM, and neutralization titers are expressed as the geometric mean with a 95% confidence interval. *p

    Journal: PLoS Neglected Tropical Diseases

    Article Title: DNA vaccination induced protective immunity against SARS CoV-2 infection in hamsterss

    doi: 10.1371/journal.pntd.0009374

    Figure Lengend Snippet: Prophylactic efficacy of SARS-CoV-2 S DNA vaccine in SARS-CoV-2-infected hamsters. (A) Time course of DNA vaccination and SARS-CoV-2 challenge. Syrian hamsters were intramuscularly immunized twice at a 3-week interval with 100 μg of control, pSARS-S or pSARS2-S, followed by electroporation. Serum samples were collected by retroorbital blood sampling at weeks 4 and 6 after the first immunization. At 4 weeks after the second immunization, Syrian hamsters were intranasally challenged with 10 5 TCID 50 SARS-CoV-2. (B) Antibodies against the SARS-CoV-2 full-length spike protein were evaluated by ELISA. (C) Vaccine-induced neutralizing activity against SARS-CoV-2 was evaluated by neutralization assay. (D) Body weight change (%) of the hamsters was recorded every day after SARS-CoV-2 challenge. Virus titers (E) and viral RNA copies (F) in the lungs of SARS-CoV-2-infected hamsters at 3 days postchallenge were determined by TCID 50 assay and qRT-PCR, respectively. Antibody titers are presented as the mean ± SEM, and neutralization titers are expressed as the geometric mean with a 95% confidence interval. *p

    Article Snippet: The proteins were then transferred to PVDF membranes and blotted with rabbit anti-Spike polyclonal antibody (40592-T62, Sino Biological).

    Techniques: Infection, Electroporation, Sampling, Enzyme-linked Immunosorbent Assay, Activity Assay, Neutralization, Quantitative RT-PCR

    Design and expression of SARS-CoV and SARS-CoV-2 spike construct variants. (A) Schematic diagram of SARS-CoV and SARS-CoV-2 spike construct variants. tPA, leader sequence from tissue-plasminogen activator; TM, transmembrane domain. (B, C) Western blot analysis of spike protein. HEK293T cells were transfected with the indicated plasmids (vector, pSARS-S, pSARS2-S, and S variants fused with tPA leader sequence). The cell lysates were collected and probed with anti-Spike antibody, and anti-β-actin antibody was used as an internal control.

    Journal: PLoS Neglected Tropical Diseases

    Article Title: DNA vaccination induced protective immunity against SARS CoV-2 infection in hamsterss

    doi: 10.1371/journal.pntd.0009374

    Figure Lengend Snippet: Design and expression of SARS-CoV and SARS-CoV-2 spike construct variants. (A) Schematic diagram of SARS-CoV and SARS-CoV-2 spike construct variants. tPA, leader sequence from tissue-plasminogen activator; TM, transmembrane domain. (B, C) Western blot analysis of spike protein. HEK293T cells were transfected with the indicated plasmids (vector, pSARS-S, pSARS2-S, and S variants fused with tPA leader sequence). The cell lysates were collected and probed with anti-Spike antibody, and anti-β-actin antibody was used as an internal control.

    Article Snippet: The proteins were then transferred to PVDF membranes and blotted with rabbit anti-Spike polyclonal antibody (40592-T62, Sino Biological).

    Techniques: Expressing, Construct, Sequencing, Western Blot, Transfection, Plasmid Preparation

    SARS-CoV-2 S DNA vaccine induced long-term humoral immunity and cross-protection against the SARS-CoV-2 with D614G mutation. BALB/c mice (n = 4 per group) were intramuscularly immunized three times at a 3-week interval with 100 μg of vector, pSARS-S or pSARS2-S, followed by electroporation. Serum samples were collected at the indicated time points after the first immunization. (A) Antibodies against the SARS-CoV-2 full-length spike protein were evaluated by ELISA. (B, C) Vaccine-induced neutralizing activity against SARS-CoV-2 with D614 or G614 genotypes was evaluated by neutralization assay. Antibody titers are presented as the mean ± SEM, and neutralization titers are expressed as the geometric mean with a 95% confidence interval. *p

    Journal: PLoS Neglected Tropical Diseases

    Article Title: DNA vaccination induced protective immunity against SARS CoV-2 infection in hamsterss

    doi: 10.1371/journal.pntd.0009374

    Figure Lengend Snippet: SARS-CoV-2 S DNA vaccine induced long-term humoral immunity and cross-protection against the SARS-CoV-2 with D614G mutation. BALB/c mice (n = 4 per group) were intramuscularly immunized three times at a 3-week interval with 100 μg of vector, pSARS-S or pSARS2-S, followed by electroporation. Serum samples were collected at the indicated time points after the first immunization. (A) Antibodies against the SARS-CoV-2 full-length spike protein were evaluated by ELISA. (B, C) Vaccine-induced neutralizing activity against SARS-CoV-2 with D614 or G614 genotypes was evaluated by neutralization assay. Antibody titers are presented as the mean ± SEM, and neutralization titers are expressed as the geometric mean with a 95% confidence interval. *p

    Article Snippet: The proteins were then transferred to PVDF membranes and blotted with rabbit anti-Spike polyclonal antibody (40592-T62, Sino Biological).

    Techniques: Mutagenesis, Mouse Assay, Plasmid Preparation, Electroporation, Enzyme-linked Immunosorbent Assay, Activity Assay, Neutralization

    Inducible Bronchus Associated Lymphoid Tissues (iBALT) formation upon MVA/S and MVA/S1 vaccination. Frozen lung sections from vaccinated mice were either stained for H E to analyze tissue structure and formation of iBALT aggregates (A), or immunofluorescence stained to visualize B cell and T cell (B) forming B cell follicle like structure (iBALT) induced by MVA/S vaccination given via i.m. route (right panel), and compared with unvaccinated control mice (left panel). Total number of iBALT like structures visualized in each section per mice was quantified and compared between the groups (C). The p value was calculated using non parametric mann-whitney test. (D) Lung immune responses in bronchoalveolar lavage (BAL) samples collected after euthanizations (three weeks post-boost) were measured using ELISA. SARS-CoV-2 S protein-specific binding IgG and IgA antibodies measured, and titters were presented in column graphs. The data represent mean responses in each group (n = 5) ± SEM.

    Journal: bioRxiv

    Article Title: Modified Vaccinia Ankara Based SARS-CoV-2 Vaccine Expressing Full-Length Spike Induces Strong Neutralizing Antibody Response

    doi: 10.1101/2020.06.27.175166

    Figure Lengend Snippet: Inducible Bronchus Associated Lymphoid Tissues (iBALT) formation upon MVA/S and MVA/S1 vaccination. Frozen lung sections from vaccinated mice were either stained for H E to analyze tissue structure and formation of iBALT aggregates (A), or immunofluorescence stained to visualize B cell and T cell (B) forming B cell follicle like structure (iBALT) induced by MVA/S vaccination given via i.m. route (right panel), and compared with unvaccinated control mice (left panel). Total number of iBALT like structures visualized in each section per mice was quantified and compared between the groups (C). The p value was calculated using non parametric mann-whitney test. (D) Lung immune responses in bronchoalveolar lavage (BAL) samples collected after euthanizations (three weeks post-boost) were measured using ELISA. SARS-CoV-2 S protein-specific binding IgG and IgA antibodies measured, and titters were presented in column graphs. The data represent mean responses in each group (n = 5) ± SEM.

    Article Snippet: Proteins were transferred to a nitrocellulose membrane, blocked with 1% casein blocker overnight (Cat#1610782 Biorad), and incubated for 1 h at room temperature with anti-SARS-CoV-2 spike mouse mAb (Cat # GTX632604, GeneTex) for MVA/S and rabbit SARS-CoV-2 RBD polyclonal antibody (Cat# 40592-T62, Sino Biological) for MVA/S1 diluted 1:2500 in blocking buffer, respectively.

    Techniques: Mouse Assay, Staining, Immunofluorescence, MANN-WHITNEY, Enzyme-linked Immunosorbent Assay, Binding Assay

    Neutralizing activity against SARS-CoV-2. (A) Percent neutralization of SARS-CoV-2 virus expressing GFP. Serum collected from the naïve animals used as negative controls. (B) Neutralization titer against SARS-CoV-2 virus expressing GFP. (C, D) Correlations between neutralization titer and ELISA binding titer.

    Journal: bioRxiv

    Article Title: Modified Vaccinia Ankara Based SARS-CoV-2 Vaccine Expressing Full-Length Spike Induces Strong Neutralizing Antibody Response

    doi: 10.1101/2020.06.27.175166

    Figure Lengend Snippet: Neutralizing activity against SARS-CoV-2. (A) Percent neutralization of SARS-CoV-2 virus expressing GFP. Serum collected from the naïve animals used as negative controls. (B) Neutralization titer against SARS-CoV-2 virus expressing GFP. (C, D) Correlations between neutralization titer and ELISA binding titer.

    Article Snippet: Proteins were transferred to a nitrocellulose membrane, blocked with 1% casein blocker overnight (Cat#1610782 Biorad), and incubated for 1 h at room temperature with anti-SARS-CoV-2 spike mouse mAb (Cat # GTX632604, GeneTex) for MVA/S and rabbit SARS-CoV-2 RBD polyclonal antibody (Cat# 40592-T62, Sino Biological) for MVA/S1 diluted 1:2500 in blocking buffer, respectively.

    Techniques: Activity Assay, Neutralization, Expressing, Enzyme-linked Immunosorbent Assay, Binding Assay

    Analyzing SARS-CoV-2 RBD and S1 proteins affinities to human ACE2 (hACE2) proteins using biolayer interferometry (BLI). (A) Bio-Layer Interferometry sensograms of the binding of SARS-CoV-2 S1 and RBD proteins to immobilized Fc-human ACE2, after incubation of the analytes at 25°C for 0and 60 minutes. The traces represent BLI response curves for SARS-CoV-2 proteins serially diluted from 800nM to 12.5nM, as indicated. Dotted lines show raw response values, while bold solid lines show the fitted trace. Association and dissociation phases were monitored for 300s and 600s, respectively. The data was globally fit using a 1:1 binding model to estimate binding affinity. (B) Binding affinity specifications of S1 and RBD proteins against hu-ACE2.

    Journal: bioRxiv

    Article Title: Modified Vaccinia Ankara Based SARS-CoV-2 Vaccine Expressing Full-Length Spike Induces Strong Neutralizing Antibody Response

    doi: 10.1101/2020.06.27.175166

    Figure Lengend Snippet: Analyzing SARS-CoV-2 RBD and S1 proteins affinities to human ACE2 (hACE2) proteins using biolayer interferometry (BLI). (A) Bio-Layer Interferometry sensograms of the binding of SARS-CoV-2 S1 and RBD proteins to immobilized Fc-human ACE2, after incubation of the analytes at 25°C for 0and 60 minutes. The traces represent BLI response curves for SARS-CoV-2 proteins serially diluted from 800nM to 12.5nM, as indicated. Dotted lines show raw response values, while bold solid lines show the fitted trace. Association and dissociation phases were monitored for 300s and 600s, respectively. The data was globally fit using a 1:1 binding model to estimate binding affinity. (B) Binding affinity specifications of S1 and RBD proteins against hu-ACE2.

    Article Snippet: Proteins were transferred to a nitrocellulose membrane, blocked with 1% casein blocker overnight (Cat#1610782 Biorad), and incubated for 1 h at room temperature with anti-SARS-CoV-2 spike mouse mAb (Cat # GTX632604, GeneTex) for MVA/S and rabbit SARS-CoV-2 RBD polyclonal antibody (Cat# 40592-T62, Sino Biological) for MVA/S1 diluted 1:2500 in blocking buffer, respectively.

    Techniques: Binding Assay, Incubation

    Antibody responses induced by MVA/S or MVA/S1 in mice. BALB/c mice were immunized on week 0 and 3 with recombinant MVAs expressing either S (MVA/S) (n=5) or S1 (MVA/S1) (n=5) in a prime-boost strategy. Unvaccinated (naïve) animals served as controls (n=5). (A) Binding IgG antibody response for individual proteins measured using ELISA at two weeks after boost. (B) Endpoint IgG titers against SARS-CoV-2 RBD, S1 and S measured at week 2 after immunization. The data show mean response in each group (n = 5) ± SEM. (C) Binding antibody response determined using Luminex assay at 3 weeks post boost. The pie graphs show the relative proportions of binding to three proteins in each group. (D) IgG subclass and soluble Fc receptor binding analysis of RBD and S1 specific IgG measured using the Luminex assay. Raw values are presented as in mean fluorescence intensity (MFI) in bar graph. The data represent mean responses in each group (n = 5) ± SEM.

    Journal: bioRxiv

    Article Title: Modified Vaccinia Ankara Based SARS-CoV-2 Vaccine Expressing Full-Length Spike Induces Strong Neutralizing Antibody Response

    doi: 10.1101/2020.06.27.175166

    Figure Lengend Snippet: Antibody responses induced by MVA/S or MVA/S1 in mice. BALB/c mice were immunized on week 0 and 3 with recombinant MVAs expressing either S (MVA/S) (n=5) or S1 (MVA/S1) (n=5) in a prime-boost strategy. Unvaccinated (naïve) animals served as controls (n=5). (A) Binding IgG antibody response for individual proteins measured using ELISA at two weeks after boost. (B) Endpoint IgG titers against SARS-CoV-2 RBD, S1 and S measured at week 2 after immunization. The data show mean response in each group (n = 5) ± SEM. (C) Binding antibody response determined using Luminex assay at 3 weeks post boost. The pie graphs show the relative proportions of binding to three proteins in each group. (D) IgG subclass and soluble Fc receptor binding analysis of RBD and S1 specific IgG measured using the Luminex assay. Raw values are presented as in mean fluorescence intensity (MFI) in bar graph. The data represent mean responses in each group (n = 5) ± SEM.

    Article Snippet: Proteins were transferred to a nitrocellulose membrane, blocked with 1% casein blocker overnight (Cat#1610782 Biorad), and incubated for 1 h at room temperature with anti-SARS-CoV-2 spike mouse mAb (Cat # GTX632604, GeneTex) for MVA/S and rabbit SARS-CoV-2 RBD polyclonal antibody (Cat# 40592-T62, Sino Biological) for MVA/S1 diluted 1:2500 in blocking buffer, respectively.

    Techniques: Mouse Assay, Recombinant, Expressing, Binding Assay, Enzyme-linked Immunosorbent Assay, Luminex, Fluorescence

    ACE2 is required for the cellular uptake and targeting of SARS-CoV-2 S protein RBD-tagged EVs. (A)Western blot analysis of human ACE2 was performed with cell lysates from various cell lines or organoid as where indicated using antibody against ACE2. (B) Schematic illustration of RBD-GFP-loaded EVs. Cultured cells were transfected with RBD-VSVG vector or empty vector (NC-vector) in combination with CD9-GFP vector. 48 h post transfection, the EV fraction that was labeled with GFP was collected(EV-RBD-GFP + /EV-NC-GFP + ). (C) Flow cytometric analysis of GFP ratio in different cell lines after incubating with 1 × 10 9 of EV-RBD-GFP+ or EV-NC-GFP+ for 6 h. Different symbols correspond to independent experiments. Data are shown as mean ± SEM of three independent experiments, each individual experiment performs triplicate. (D, E and F) Representative images of Caco-2 cells (D), iPS cells-derived cardiomyocytes (E) and intestinal organoids (F) after application of 1 × 10 9 EV-RBD-GFP + or EV-NC-GFP + for 6 h. The GFP ratio was determined by flow cytometric analysis. (G) The binding affinity for RBD-tagged EVs with ACE2 protein was quantified by ELISA. Different concentration of EVs were incubated and followed by the anti-CD9 as the detector antibody. Each concentration was repeated with six wells. Data are shown as mean ± SEM of three independent experiments. (H and I) Representative images of ACE2-A549 cells showed an earlier plasma membrane distribution of EV-RBD-GFP + or EV-NC-GFP + incubation for 1 h (H) and 6 h (I) with or without pre-treated with ACE2 antibody. Fluorescence intensity of GFP was analyzed by ImageJ software were averaged from six different fields for each. Data are shown as mean ± SEM of three independent experiments. ** p

    Journal: Journal of Controlled Release

    Article Title: Tagged extracellular vesicles with the RBD of the viral spike protein for delivery of antiviral agents against SARS-COV-2 infection

    doi: 10.1016/j.jconrel.2021.05.049

    Figure Lengend Snippet: ACE2 is required for the cellular uptake and targeting of SARS-CoV-2 S protein RBD-tagged EVs. (A)Western blot analysis of human ACE2 was performed with cell lysates from various cell lines or organoid as where indicated using antibody against ACE2. (B) Schematic illustration of RBD-GFP-loaded EVs. Cultured cells were transfected with RBD-VSVG vector or empty vector (NC-vector) in combination with CD9-GFP vector. 48 h post transfection, the EV fraction that was labeled with GFP was collected(EV-RBD-GFP + /EV-NC-GFP + ). (C) Flow cytometric analysis of GFP ratio in different cell lines after incubating with 1 × 10 9 of EV-RBD-GFP+ or EV-NC-GFP+ for 6 h. Different symbols correspond to independent experiments. Data are shown as mean ± SEM of three independent experiments, each individual experiment performs triplicate. (D, E and F) Representative images of Caco-2 cells (D), iPS cells-derived cardiomyocytes (E) and intestinal organoids (F) after application of 1 × 10 9 EV-RBD-GFP + or EV-NC-GFP + for 6 h. The GFP ratio was determined by flow cytometric analysis. (G) The binding affinity for RBD-tagged EVs with ACE2 protein was quantified by ELISA. Different concentration of EVs were incubated and followed by the anti-CD9 as the detector antibody. Each concentration was repeated with six wells. Data are shown as mean ± SEM of three independent experiments. (H and I) Representative images of ACE2-A549 cells showed an earlier plasma membrane distribution of EV-RBD-GFP + or EV-NC-GFP + incubation for 1 h (H) and 6 h (I) with or without pre-treated with ACE2 antibody. Fluorescence intensity of GFP was analyzed by ImageJ software were averaged from six different fields for each. Data are shown as mean ± SEM of three independent experiments. ** p

    Article Snippet: 2.5 Western blotCells, purified EVs or tissues were lysed with RIPA buffer (Santa Cruz,USA) and cleared lysate was collected by centrifugation for protein separation on SDS-polyacrylamide gel(10%), followed by transfer onto PVDF membranes(Millipore), blocked with 5% nonfat dry milk in TBST and reacted with primary antibodies recognizing FLAG M2 tag(Sigma,1:2000), SARS-CoV-2 RBD(Sinobiological,1:500), CD9(Abcam,1:2000), Alix(Cell Signaling Technology,1:1000), GM130(Abcam,1:1000), Calnexin(Abcam,1:1000), hACE2(Abcam,# ab108209,only react with human species,1:2000) and GAPDH(Santa Cruz,1:2000).

    Techniques: Cell Culture, Transfection, Plasmid Preparation, Labeling, Derivative Assay, Binding Assay, Enzyme-linked Immunosorbent Assay, Concentration Assay, Incubation, Fluorescence, Software

    Characterization of physical properties of SARS-CoV-2 S protein RBD-tagged EVs. (A) The ratio of particles to protein for each sample is shown. (B)Histogram showing the size distribution of the purified EVs as analyzed by NTA. (C) TEM or SEM images of RBD-tagged or control EVs purified from 293T cells. (D and E) EV protein and RNA concentration measurement. EV protein or RNA was extracted from 1 × 10 8 EVs and protein concentration was measured by BCA protein quantification (D) and RNA concentration was determined by NanoDrop (E). Data are shown as mean ± SEM of four independent experiments, each experiment performs triplicate.

    Journal: Journal of Controlled Release

    Article Title: Tagged extracellular vesicles with the RBD of the viral spike protein for delivery of antiviral agents against SARS-COV-2 infection

    doi: 10.1016/j.jconrel.2021.05.049

    Figure Lengend Snippet: Characterization of physical properties of SARS-CoV-2 S protein RBD-tagged EVs. (A) The ratio of particles to protein for each sample is shown. (B)Histogram showing the size distribution of the purified EVs as analyzed by NTA. (C) TEM or SEM images of RBD-tagged or control EVs purified from 293T cells. (D and E) EV protein and RNA concentration measurement. EV protein or RNA was extracted from 1 × 10 8 EVs and protein concentration was measured by BCA protein quantification (D) and RNA concentration was determined by NanoDrop (E). Data are shown as mean ± SEM of four independent experiments, each experiment performs triplicate.

    Article Snippet: 2.5 Western blotCells, purified EVs or tissues were lysed with RIPA buffer (Santa Cruz,USA) and cleared lysate was collected by centrifugation for protein separation on SDS-polyacrylamide gel(10%), followed by transfer onto PVDF membranes(Millipore), blocked with 5% nonfat dry milk in TBST and reacted with primary antibodies recognizing FLAG M2 tag(Sigma,1:2000), SARS-CoV-2 RBD(Sinobiological,1:500), CD9(Abcam,1:2000), Alix(Cell Signaling Technology,1:1000), GM130(Abcam,1:1000), Calnexin(Abcam,1:1000), hACE2(Abcam,# ab108209,only react with human species,1:2000) and GAPDH(Santa Cruz,1:2000).

    Techniques: Purification, Transmission Electron Microscopy, Concentration Assay, Protein Concentration

    SARS-CoV-2 S protein RBD-tagged EVs specifically target tissues in vivo . (A) Western blot analysis of tissue distribution of hACE2 in humanized mice using antibody against human ACE2. (B,C,D) After intravenous injection of DiD-labeled EV-RBD or EV-NC in hACE2 mice( n = 4), the red-fluorescence of whole animal was detected at the indicated time points by IVIS spectrum (B) Representative IVIS images of various organs were acquired at the indicated time points through intravenous injection of EVs(C). Radiant efficiency was measured using Living Image software. Four mice were sacrificed in each time-point for tissue collection(D). (E, F, G) Serum cytokines concentrations of IL-6 (E), TNF-α (F) and IFN-β (G) were measured by ELISA. Serum samples were from hACE2 mice after intravenous injection of EV-RBD or EV-NC at 24 h. Each mouse serum was repeated with four wells. All data expressed as mean ± SEM of four independent experiments. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

    Journal: Journal of Controlled Release

    Article Title: Tagged extracellular vesicles with the RBD of the viral spike protein for delivery of antiviral agents against SARS-COV-2 infection

    doi: 10.1016/j.jconrel.2021.05.049

    Figure Lengend Snippet: SARS-CoV-2 S protein RBD-tagged EVs specifically target tissues in vivo . (A) Western blot analysis of tissue distribution of hACE2 in humanized mice using antibody against human ACE2. (B,C,D) After intravenous injection of DiD-labeled EV-RBD or EV-NC in hACE2 mice( n = 4), the red-fluorescence of whole animal was detected at the indicated time points by IVIS spectrum (B) Representative IVIS images of various organs were acquired at the indicated time points through intravenous injection of EVs(C). Radiant efficiency was measured using Living Image software. Four mice were sacrificed in each time-point for tissue collection(D). (E, F, G) Serum cytokines concentrations of IL-6 (E), TNF-α (F) and IFN-β (G) were measured by ELISA. Serum samples were from hACE2 mice after intravenous injection of EV-RBD or EV-NC at 24 h. Each mouse serum was repeated with four wells. All data expressed as mean ± SEM of four independent experiments. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

    Article Snippet: 2.5 Western blotCells, purified EVs or tissues were lysed with RIPA buffer (Santa Cruz,USA) and cleared lysate was collected by centrifugation for protein separation on SDS-polyacrylamide gel(10%), followed by transfer onto PVDF membranes(Millipore), blocked with 5% nonfat dry milk in TBST and reacted with primary antibodies recognizing FLAG M2 tag(Sigma,1:2000), SARS-CoV-2 RBD(Sinobiological,1:500), CD9(Abcam,1:2000), Alix(Cell Signaling Technology,1:1000), GM130(Abcam,1:1000), Calnexin(Abcam,1:1000), hACE2(Abcam,# ab108209,only react with human species,1:2000) and GAPDH(Santa Cruz,1:2000).

    Techniques: In Vivo, Western Blot, Mouse Assay, Injection, Labeling, Fluorescence, Software, Enzyme-linked Immunosorbent Assay

    Delivery of siRNAs as a model system to identify inhibition of SARS-CoV-2 pseudovirus infection in vivo . (A) The efficiency of siRNA loading in EVs after electroporation as measured by real-time PCR. Each sample contained mixture of 150 μg siRNA-GFP and 150 μg EVs or 150 μg naked siRNA. After electroporation, purified EVs by ultracentrifugation to remove unencapsulated siRNA and each EV pellet treated with or without Benzonase. All electroporation samples were prepared in triplicate and each RNA isolate was analyzed in duplicate. (B) Scheme of the possibility of loading unmodified or modified EVs with specific siRNA into target cells. (C) Representative images of pseudotyped SARS-CoV-2-GFP-infected ACE2-A549 cells after incubation with different origin of EVs. The ACE2-A549 cells was infected with SARS-CoV-2-GFP for 24 h(MOI = 2), and then incubated for 24 h with naked si-GFP, EV-NC or EV-RBD electroporated with the siRNA, respectively. The GFP ratio was determined by flow cytometric analysis. Data are shown as mean ± SEM of three independent experiments in triplicates. (D) hACE2 mice(n = 4) were inoculated intranasally with pseudotyped SARS-CoV-2-GFP at a multiplicity of infection (MOI) of 50 per 50 μl inoculum volume per mouse. At 24 h post-infection,150 μg RBD-tagged EVs(EV-RBD-si-GFP) or control EVs (EV-NC-si-GFP) loaded with 150 μg GFP siRNA were injected into tail veins, and all mice were sacrificed at 48 h post injection for lung tissues collection.(E) Immunofluorescence staining of mouse lung paraffin sections for pseudotyped SARS-CoV-2-GFP (green, white arrows) and DAPI(blue). (F) Each hACE2 mouse(n = 4) injected with 150 μg RBD-tagged EVs encapsulated GFP siRNA or control EVs in 24 h and later inoculated intranasally with pseudotyped SARS-CoV-2-GFP at MOI of 50. All mice were sacrificed at 48 h post infection for lung tissues collection (G) Immunofluorescence staining analysis for pseudotyped SARS-CoV-2-GFP infected cells in lung paraffin sections (green, white arrows). Fluorescence intensity of GFP in the regions were analyzed through ImageJ software. Data are shown as mean ± SEM of four independent experiments in triplicates.**p

    Journal: Journal of Controlled Release

    Article Title: Tagged extracellular vesicles with the RBD of the viral spike protein for delivery of antiviral agents against SARS-COV-2 infection

    doi: 10.1016/j.jconrel.2021.05.049

    Figure Lengend Snippet: Delivery of siRNAs as a model system to identify inhibition of SARS-CoV-2 pseudovirus infection in vivo . (A) The efficiency of siRNA loading in EVs after electroporation as measured by real-time PCR. Each sample contained mixture of 150 μg siRNA-GFP and 150 μg EVs or 150 μg naked siRNA. After electroporation, purified EVs by ultracentrifugation to remove unencapsulated siRNA and each EV pellet treated with or without Benzonase. All electroporation samples were prepared in triplicate and each RNA isolate was analyzed in duplicate. (B) Scheme of the possibility of loading unmodified or modified EVs with specific siRNA into target cells. (C) Representative images of pseudotyped SARS-CoV-2-GFP-infected ACE2-A549 cells after incubation with different origin of EVs. The ACE2-A549 cells was infected with SARS-CoV-2-GFP for 24 h(MOI = 2), and then incubated for 24 h with naked si-GFP, EV-NC or EV-RBD electroporated with the siRNA, respectively. The GFP ratio was determined by flow cytometric analysis. Data are shown as mean ± SEM of three independent experiments in triplicates. (D) hACE2 mice(n = 4) were inoculated intranasally with pseudotyped SARS-CoV-2-GFP at a multiplicity of infection (MOI) of 50 per 50 μl inoculum volume per mouse. At 24 h post-infection,150 μg RBD-tagged EVs(EV-RBD-si-GFP) or control EVs (EV-NC-si-GFP) loaded with 150 μg GFP siRNA were injected into tail veins, and all mice were sacrificed at 48 h post injection for lung tissues collection.(E) Immunofluorescence staining of mouse lung paraffin sections for pseudotyped SARS-CoV-2-GFP (green, white arrows) and DAPI(blue). (F) Each hACE2 mouse(n = 4) injected with 150 μg RBD-tagged EVs encapsulated GFP siRNA or control EVs in 24 h and later inoculated intranasally with pseudotyped SARS-CoV-2-GFP at MOI of 50. All mice were sacrificed at 48 h post infection for lung tissues collection (G) Immunofluorescence staining analysis for pseudotyped SARS-CoV-2-GFP infected cells in lung paraffin sections (green, white arrows). Fluorescence intensity of GFP in the regions were analyzed through ImageJ software. Data are shown as mean ± SEM of four independent experiments in triplicates.**p

    Article Snippet: 2.5 Western blotCells, purified EVs or tissues were lysed with RIPA buffer (Santa Cruz,USA) and cleared lysate was collected by centrifugation for protein separation on SDS-polyacrylamide gel(10%), followed by transfer onto PVDF membranes(Millipore), blocked with 5% nonfat dry milk in TBST and reacted with primary antibodies recognizing FLAG M2 tag(Sigma,1:2000), SARS-CoV-2 RBD(Sinobiological,1:500), CD9(Abcam,1:2000), Alix(Cell Signaling Technology,1:1000), GM130(Abcam,1:1000), Calnexin(Abcam,1:1000), hACE2(Abcam,# ab108209,only react with human species,1:2000) and GAPDH(Santa Cruz,1:2000).

    Techniques: Inhibition, Infection, In Vivo, Electroporation, Real-time Polymerase Chain Reaction, Purification, Modification, Incubation, Mouse Assay, Injection, Immunofluorescence, Staining, Fluorescence, Software

    Construction of SARS-CoV-2 S protein RBD-tagged EVs. (A) Detection of SARS-CoV-2 Spike protein in cell lysates or EV pellets by western blot. 293T cells were transfected with plasmid expressing the FLAG-tagged Spike protein. EVs pelleted from the media were analyzed by western blot for the presence of the Spike protein using the FLAG M2 antibody. Purified EVs were characterized for EV markers CD9 and Alix, and non-EV markers calnexin (endoplasmic reticulum marker) and GM130 (Golgi matrix marker). (B) Schematic illustration of RBD-VSVG fusion-loaded EVs. The VSVG ectodomain was replaced by the RBD domain of SARS-CoV-2 (right, red), followed by a transmembrane helix and a cytoplasmic tail of VSVG (right, green). (C) Western blot analysis of EVs purified from 293T cells transfected with or without RBD-VSVG-vector for 48 h. CD9 or Alix proteins served as markers for the presence of EVs.(D, E) Schematic presentation of affinity pull-down of EVs by anti-RBD coupled magnetic beads. (D)EVs isolated from supernatants of transfected or non-transfected 293T cells were subjected to anti-RBD pull-down assay, then total protein of EVs was analyzed by western blot (E). (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

    Journal: Journal of Controlled Release

    Article Title: Tagged extracellular vesicles with the RBD of the viral spike protein for delivery of antiviral agents against SARS-COV-2 infection

    doi: 10.1016/j.jconrel.2021.05.049

    Figure Lengend Snippet: Construction of SARS-CoV-2 S protein RBD-tagged EVs. (A) Detection of SARS-CoV-2 Spike protein in cell lysates or EV pellets by western blot. 293T cells were transfected with plasmid expressing the FLAG-tagged Spike protein. EVs pelleted from the media were analyzed by western blot for the presence of the Spike protein using the FLAG M2 antibody. Purified EVs were characterized for EV markers CD9 and Alix, and non-EV markers calnexin (endoplasmic reticulum marker) and GM130 (Golgi matrix marker). (B) Schematic illustration of RBD-VSVG fusion-loaded EVs. The VSVG ectodomain was replaced by the RBD domain of SARS-CoV-2 (right, red), followed by a transmembrane helix and a cytoplasmic tail of VSVG (right, green). (C) Western blot analysis of EVs purified from 293T cells transfected with or without RBD-VSVG-vector for 48 h. CD9 or Alix proteins served as markers for the presence of EVs.(D, E) Schematic presentation of affinity pull-down of EVs by anti-RBD coupled magnetic beads. (D)EVs isolated from supernatants of transfected or non-transfected 293T cells were subjected to anti-RBD pull-down assay, then total protein of EVs was analyzed by western blot (E). (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

    Article Snippet: 2.5 Western blotCells, purified EVs or tissues were lysed with RIPA buffer (Santa Cruz,USA) and cleared lysate was collected by centrifugation for protein separation on SDS-polyacrylamide gel(10%), followed by transfer onto PVDF membranes(Millipore), blocked with 5% nonfat dry milk in TBST and reacted with primary antibodies recognizing FLAG M2 tag(Sigma,1:2000), SARS-CoV-2 RBD(Sinobiological,1:500), CD9(Abcam,1:2000), Alix(Cell Signaling Technology,1:1000), GM130(Abcam,1:1000), Calnexin(Abcam,1:1000), hACE2(Abcam,# ab108209,only react with human species,1:2000) and GAPDH(Santa Cruz,1:2000).

    Techniques: Western Blot, Transfection, Plasmid Preparation, Expressing, Purification, Marker, Magnetic Beads, Isolation, Pull Down Assay