insect cells  (Sino Biological)


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    Name:
    Influenza A H5N1 Hemagglutinin HA Insect Cells Cell Lysate
    Description:
    Baculovirus Insect Cells that Influenza A H5N1 A Vietnam 1194 2004 Hemagglutinin HA Heterodimer transfected overexpressed for Western blot WB positive control The whole cell lysate is provided in 1X Sample Buffer 1X modified RIPA buffer 1X SDS loading buffer
    Catalog Number:
    11062-V08BL
    Price:
    195
    Category:
    Lysate
    Size:
    300µg
    Cell Type:
    Baculovirus-Insect Cells
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    Structured Review

    Sino Biological insect cells
    Baculovirus Insect Cells that Influenza A H5N1 A Vietnam 1194 2004 Hemagglutinin HA Heterodimer transfected overexpressed for Western blot WB positive control The whole cell lysate is provided in 1X Sample Buffer 1X modified RIPA buffer 1X SDS loading buffer
    https://www.bioz.com/result/insect cells/product/Sino Biological
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    insect cells - by Bioz Stars, 2021-07
    86/100 stars

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    Related Articles

    Expressing:

    Article Title: The crystal structure of Ac-AChBP in complex with α-conotoxin LvIA reveals the mechanism of its selectivity towards different nAChR subtypes
    Article Snippet: The masses of the peptides were verified by matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry. .. Protein expression and purification Ac-AChBP was overexpressed in High Five insect cells maintained in SIM-HF medium (Sino Biological Inc., China) using the Bac-to-Bac baculovirus expression system (Invitrogen, Thermo Fisher Scientific, USA). .. Sf9 insect cells were maintained in Insect-XPRESS™ Protein-free Insect Cell Medium (Lonza, Switzerland).

    Purification:

    Article Title: The crystal structure of Ac-AChBP in complex with α-conotoxin LvIA reveals the mechanism of its selectivity towards different nAChR subtypes
    Article Snippet: The masses of the peptides were verified by matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry. .. Protein expression and purification Ac-AChBP was overexpressed in High Five insect cells maintained in SIM-HF medium (Sino Biological Inc., China) using the Bac-to-Bac baculovirus expression system (Invitrogen, Thermo Fisher Scientific, USA). .. Sf9 insect cells were maintained in Insect-XPRESS™ Protein-free Insect Cell Medium (Lonza, Switzerland).

    Article Title: Modulation of LSD1 phosphorylation by CK2/WIP1 regulates RNF168-dependent 53BP1 recruitment in response to DNA damage
    Article Snippet: Bleomycin was purchased from Nippon Kayaku Co, etoposide (E1383) and TBB (T0826) were from Sigma, respectively. .. GST-CK2α (10630-H09B) and His-GST CK2α’ (11018-H20B) produced in and purified from insect cells were purchased from Sino Biological Inc. Rabbit polyclonal antibodies used in this study including anti-HA (A190–208A), anti-MYC (A190–205A), anti-LSD1 (A300–215A), anti-53BP1 (A300–272A), anti-CK2α (A300–198A), anti-CK2α’ (A300–199A), anti-CK2β (A301–984A), anti-GAPDH (A300–643A) and anti-WIP1 (A300–664A) were purchased from Bethyl, while rabbit anti-RNF168 (06–1130) polyclonal antibodies were from Millipore. .. Mouse monoclonal antibodies including anti-FLAG M2 (F1804) and anti-β-actin (A5441) were from Sigma.

    Article Title: A single dose of an adenovirus-vectored vaccine provides protection against SARS-CoV-2 challenge
    Article Snippet: .. ELISAFor SARS-CoV-2 S-specific IgG assays in mice, 96-well polystyrene high-binding microplates (Corning, USA) were coated with 2 μg/mL recombinant SARS-CoV-2 S protein purified from insect cells (Sino Biological, China) in carbonate-bicarbonate buffer pH 9.6, and the plates were incubated at 4 °C overnight. .. The plates were then blocked at 37 °C for 1 h with PBS pH 7.4 in 5% skim milk (blocking buffer) and washed with PBST.

    BAC Assay:

    Article Title: The crystal structure of Ac-AChBP in complex with α-conotoxin LvIA reveals the mechanism of its selectivity towards different nAChR subtypes
    Article Snippet: The masses of the peptides were verified by matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry. .. Protein expression and purification Ac-AChBP was overexpressed in High Five insect cells maintained in SIM-HF medium (Sino Biological Inc., China) using the Bac-to-Bac baculovirus expression system (Invitrogen, Thermo Fisher Scientific, USA). .. Sf9 insect cells were maintained in Insect-XPRESS™ Protein-free Insect Cell Medium (Lonza, Switzerland).

    Isolation:

    Article Title: Molecular Adjuvant Ag85A Enhances Protection against Influenza A Virus in Mice Following DNA Vaccination
    Article Snippet: .. Cells were isolated, washed, adjusted to a concentration of 5 × 106 cells per mL, and grown in a 96-well flat-bottom plate (5 × 105 cells per well) in Roswell Park Memorial Institute medium (RPMI) 1640 medium, supplemented with HEPES (N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid), glutamine and 10% heat-inactivated fetal calf serum (FCS). .. Cells were stimulated separately with 0.5 µg concanavalin A (ConA) or 1.25 µg recombinant HA of A/PR/8/34 (catalog number: 11684-V08H, Sino Biological Inc., Beijing, China) and incubated at 37 °C in a humidified CO2 incubator.

    Concentration Assay:

    Article Title: Molecular Adjuvant Ag85A Enhances Protection against Influenza A Virus in Mice Following DNA Vaccination
    Article Snippet: .. Cells were isolated, washed, adjusted to a concentration of 5 × 106 cells per mL, and grown in a 96-well flat-bottom plate (5 × 105 cells per well) in Roswell Park Memorial Institute medium (RPMI) 1640 medium, supplemented with HEPES (N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid), glutamine and 10% heat-inactivated fetal calf serum (FCS). .. Cells were stimulated separately with 0.5 µg concanavalin A (ConA) or 1.25 µg recombinant HA of A/PR/8/34 (catalog number: 11684-V08H, Sino Biological Inc., Beijing, China) and incubated at 37 °C in a humidified CO2 incubator.

    In Vitro:

    Article Title: Recombinant SARS-CoV-2 RBD with a built in T helper epitope induces strong neutralization antibody response
    Article Snippet: After diluted with normal saline, S1-4 and S1-5 protein were mixed respectively with aluminum adjuvant and adjusted antigen concentration in the final adsorption product was 800 μg/mL. .. 3.2 S1-4 protein possessed antigenicity in vitro The antigenicity of S1-4 protein was demonstrated by WB analysis with a commercial recombinant RBD protein (Val16 - Arg685 of spike protein; 76.45 kDa) expressed in baculovirus-insect cells by Sino Biological (Beijing, China) as positive control ( ). .. Ten micrograms of S1-4 protein and commercial recombinant RBD protein were loaded onto the first three gels ( A – 4C), with the exception of 30 µg of the two proteins in the fourth gel ( D).

    Western Blot:

    Article Title: Recombinant SARS-CoV-2 RBD with a built in T helper epitope induces strong neutralization antibody response
    Article Snippet: After diluted with normal saline, S1-4 and S1-5 protein were mixed respectively with aluminum adjuvant and adjusted antigen concentration in the final adsorption product was 800 μg/mL. .. 3.2 S1-4 protein possessed antigenicity in vitro The antigenicity of S1-4 protein was demonstrated by WB analysis with a commercial recombinant RBD protein (Val16 - Arg685 of spike protein; 76.45 kDa) expressed in baculovirus-insect cells by Sino Biological (Beijing, China) as positive control ( ). .. Ten micrograms of S1-4 protein and commercial recombinant RBD protein were loaded onto the first three gels ( A – 4C), with the exception of 30 µg of the two proteins in the fourth gel ( D).

    Recombinant:

    Article Title: Recombinant SARS-CoV-2 RBD with a built in T helper epitope induces strong neutralization antibody response
    Article Snippet: After diluted with normal saline, S1-4 and S1-5 protein were mixed respectively with aluminum adjuvant and adjusted antigen concentration in the final adsorption product was 800 μg/mL. .. 3.2 S1-4 protein possessed antigenicity in vitro The antigenicity of S1-4 protein was demonstrated by WB analysis with a commercial recombinant RBD protein (Val16 - Arg685 of spike protein; 76.45 kDa) expressed in baculovirus-insect cells by Sino Biological (Beijing, China) as positive control ( ). .. Ten micrograms of S1-4 protein and commercial recombinant RBD protein were loaded onto the first three gels ( A – 4C), with the exception of 30 µg of the two proteins in the fourth gel ( D).

    Article Title: Green Tea Catechin-Inactivated Viral Vaccine Platform
    Article Snippet: The digested proteins (0.1 mg/mL) were incubated with various concentrations of GT (0–1,000 μg) for 6 h and then subjected to SDS-PAGE to monitor mobility changes of the proteins. .. Seven different recombinant HA proteins expressed in baculovirus-insect cells were purchased from Sino Biologicals (China). .. Mass Spectrometry Analysis The HA proteins incubated with EGCG were subjected to SDS-PAGE and analyzed by liquid chromatography-MS/MS.

    Article Title: A single dose of an adenovirus-vectored vaccine provides protection against SARS-CoV-2 challenge
    Article Snippet: .. ELISAFor SARS-CoV-2 S-specific IgG assays in mice, 96-well polystyrene high-binding microplates (Corning, USA) were coated with 2 μg/mL recombinant SARS-CoV-2 S protein purified from insect cells (Sino Biological, China) in carbonate-bicarbonate buffer pH 9.6, and the plates were incubated at 4 °C overnight. .. The plates were then blocked at 37 °C for 1 h with PBS pH 7.4 in 5% skim milk (blocking buffer) and washed with PBST.

    Article Title: Site-specific N-glycosylation Characterization of Recombinant SARS-CoV-2 Spike Proteins
    Article Snippet: The zwitterionic hydrophilic interaction liquid chromatography (Zic-HILIC) materials were obtained from Fresh Bioscience (Shanghai, China). .. The recombinant SARS-CoV-2 S protein (S1+S2 ECD and RBD) expressed in insect cells (High Five) and S protein (S1 and RBD) expressed in human embryonic kidney cells (HEK293) were purchased from Sino Biological (Beijing, China). .. Codon-optimized DNA sequences encoding the SARS-CoV-2 S protein subunits were cloned into pCMV3-C-His and a baculovirus vector with a polyhistidine tag at the C-terminus for recombinant expression of these proteins in human and insect cells, respectively.

    Positive Control:

    Article Title: Recombinant SARS-CoV-2 RBD with a built in T helper epitope induces strong neutralization antibody response
    Article Snippet: After diluted with normal saline, S1-4 and S1-5 protein were mixed respectively with aluminum adjuvant and adjusted antigen concentration in the final adsorption product was 800 μg/mL. .. 3.2 S1-4 protein possessed antigenicity in vitro The antigenicity of S1-4 protein was demonstrated by WB analysis with a commercial recombinant RBD protein (Val16 - Arg685 of spike protein; 76.45 kDa) expressed in baculovirus-insect cells by Sino Biological (Beijing, China) as positive control ( ). .. Ten micrograms of S1-4 protein and commercial recombinant RBD protein were loaded onto the first three gels ( A – 4C), with the exception of 30 µg of the two proteins in the fourth gel ( D).

    Produced:

    Article Title: Modulation of LSD1 phosphorylation by CK2/WIP1 regulates RNF168-dependent 53BP1 recruitment in response to DNA damage
    Article Snippet: Bleomycin was purchased from Nippon Kayaku Co, etoposide (E1383) and TBB (T0826) were from Sigma, respectively. .. GST-CK2α (10630-H09B) and His-GST CK2α’ (11018-H20B) produced in and purified from insect cells were purchased from Sino Biological Inc. Rabbit polyclonal antibodies used in this study including anti-HA (A190–208A), anti-MYC (A190–205A), anti-LSD1 (A300–215A), anti-53BP1 (A300–272A), anti-CK2α (A300–198A), anti-CK2α’ (A300–199A), anti-CK2β (A301–984A), anti-GAPDH (A300–643A) and anti-WIP1 (A300–664A) were purchased from Bethyl, while rabbit anti-RNF168 (06–1130) polyclonal antibodies were from Millipore. .. Mouse monoclonal antibodies including anti-FLAG M2 (F1804) and anti-β-actin (A5441) were from Sigma.

    Mouse Assay:

    Article Title: A single dose of an adenovirus-vectored vaccine provides protection against SARS-CoV-2 challenge
    Article Snippet: .. ELISAFor SARS-CoV-2 S-specific IgG assays in mice, 96-well polystyrene high-binding microplates (Corning, USA) were coated with 2 μg/mL recombinant SARS-CoV-2 S protein purified from insect cells (Sino Biological, China) in carbonate-bicarbonate buffer pH 9.6, and the plates were incubated at 4 °C overnight. .. The plates were then blocked at 37 °C for 1 h with PBS pH 7.4 in 5% skim milk (blocking buffer) and washed with PBST.

    Incubation:

    Article Title: A single dose of an adenovirus-vectored vaccine provides protection against SARS-CoV-2 challenge
    Article Snippet: .. ELISAFor SARS-CoV-2 S-specific IgG assays in mice, 96-well polystyrene high-binding microplates (Corning, USA) were coated with 2 μg/mL recombinant SARS-CoV-2 S protein purified from insect cells (Sino Biological, China) in carbonate-bicarbonate buffer pH 9.6, and the plates were incubated at 4 °C overnight. .. The plates were then blocked at 37 °C for 1 h with PBS pH 7.4 in 5% skim milk (blocking buffer) and washed with PBST.

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  • 94
    Sino Biological sars cov s1
    S1 protein detection. (A) Illustration of the assay mechanism. The sample-to-answer time of this assay is 20 minutes. (B) Entire dynamic ranges of <t>SARS-CoV-2</t> S1 protein (red squares) and SARS-CoV S1 protein (black circles) in 10 times diluted human serum. The averaged background is subtracted from all data points. The solid lines are the linear fit of the data in the log-log scale. The grey shaded area marks 3×standard deviation of the background. The lower limit of detection (LLOD) for SARS-CoV-2 S1 protein and SARS-CoV S1 is 0.4 ng/mL and 0.2 ng/mL, respectively. (C) Calibration curves for S1 proteins between 0.78 and 200 ng/mL. The error bars are generated from duplicate measurements.
    Sars Cov S1, supplied by Sino Biological, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/sars cov s1/product/Sino Biological
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    sars cov s1 - by Bioz Stars, 2021-07
    94/100 stars
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    96
    Sino Biological sars cov 2
    Diagnostic accuracy parameters of both Ag-RDTs for different definitions of RT-PCR test positivity based on viral load cut-offs, where a positive RT-PCR test with a viral load below the viral load cut-off threshold is considered a negative RT-PCR test result. Points highlighted in red indicate a viral load cut-off of 5.2 log10 E gene copies/mL, which was considered the viral load cut-off for infectiousness as determined by viral culture. BD = BD Veritor™ System for Rapid Detection of <t>SARS-CoV-2</t> Ag-RDT; SD-B = Roche/SD Biosensor SARS-CoV-2 Rapid Antigen Test; PPV = positive predictive value; NPV = negative predictive value.
    Sars Cov 2, supplied by Sino Biological, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/sars cov 2/product/Sino Biological
    Average 96 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    sars cov 2 - by Bioz Stars, 2021-07
    96/100 stars
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    92
    Sino Biological tlr4
    DENV NS1 binds to platelets and induces activation through <t>TLR4</t> signal transduction. (A) The protein expression levels of TLR4 and β actin, an internal control, in human-isolated platelets from 3 different donors were detected using Western blotting (50 μg protein/lane). (B) Human-isolated platelets were preincubated with αTLR4 or a control Rabbit IgG for 1 h, and the binding of NS1 on platelet surfaces was determined by flow cytometry using FITC-conjugated anti-NS1 monoclonal antibodies (33D2-FITC) (n = 3). (C) The binding of NS1 on platelets isolated from wild-type or TLR4 knockout mice was determined by flow cytometry (n = 4). Platelets were preincubated with or without different concentrations of (D) αTLR4 (E) the TLR4 antagonist LPS-Rs, (F) the TLR4 signaling inhibitor TAK242, or the LPS inhibitor polymyxin B (10 μg/ml) for 30 min, followed by BSA or NS1 (10 μg/ml) stimulation for 1 h (n = 4). The percent fluorescence of NS1 binding and the P-selectin surface expression on platelets were analyzed by FACSCalibur flow cytometry. *P
    Tlr4, supplied by Sino Biological, used in various techniques. Bioz Stars score: 92/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/tlr4/product/Sino Biological
    Average 92 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    tlr4 - by Bioz Stars, 2021-07
    92/100 stars
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    N/A
    Baculovirus Insect Cells that Zika virus ZIKV strain Zika SPH2015 ZIKV NS5 Heterodimer transfected overexpressed for Western blot WB positive control The whole cell lysate is provided in 1X Sample
      Buy from Supplier

    Image Search Results


    S1 protein detection. (A) Illustration of the assay mechanism. The sample-to-answer time of this assay is 20 minutes. (B) Entire dynamic ranges of SARS-CoV-2 S1 protein (red squares) and SARS-CoV S1 protein (black circles) in 10 times diluted human serum. The averaged background is subtracted from all data points. The solid lines are the linear fit of the data in the log-log scale. The grey shaded area marks 3×standard deviation of the background. The lower limit of detection (LLOD) for SARS-CoV-2 S1 protein and SARS-CoV S1 is 0.4 ng/mL and 0.2 ng/mL, respectively. (C) Calibration curves for S1 proteins between 0.78 and 200 ng/mL. The error bars are generated from duplicate measurements.

    Journal: bioRxiv

    Article Title: Rapid and quantitative detection of COVID-19 markers in micro-liter sized samples

    doi: 10.1101/2020.04.20.052233

    Figure Lengend Snippet: S1 protein detection. (A) Illustration of the assay mechanism. The sample-to-answer time of this assay is 20 minutes. (B) Entire dynamic ranges of SARS-CoV-2 S1 protein (red squares) and SARS-CoV S1 protein (black circles) in 10 times diluted human serum. The averaged background is subtracted from all data points. The solid lines are the linear fit of the data in the log-log scale. The grey shaded area marks 3×standard deviation of the background. The lower limit of detection (LLOD) for SARS-CoV-2 S1 protein and SARS-CoV S1 is 0.4 ng/mL and 0.2 ng/mL, respectively. (C) Calibration curves for S1 proteins between 0.78 and 200 ng/mL. The error bars are generated from duplicate measurements.

    Article Snippet: In addition, the pattern of calibration curves with SARS-CoV-2 S1 is significantly different from that with SARS-CoV S1.

    Techniques: Generated

    Diagnostic accuracy parameters of both Ag-RDTs for different definitions of RT-PCR test positivity based on viral load cut-offs, where a positive RT-PCR test with a viral load below the viral load cut-off threshold is considered a negative RT-PCR test result. Points highlighted in red indicate a viral load cut-off of 5.2 log10 E gene copies/mL, which was considered the viral load cut-off for infectiousness as determined by viral culture. BD = BD Veritor™ System for Rapid Detection of SARS-CoV-2 Ag-RDT; SD-B = Roche/SD Biosensor SARS-CoV-2 Rapid Antigen Test; PPV = positive predictive value; NPV = negative predictive value.

    Journal: medRxiv

    Article Title: Diagnostic accuracy of rapid antigen tests in pre-/asymptomatic close contacts of individuals with a confirmed SARS-CoV-2 infection

    doi: 10.1101/2021.03.18.21253874

    Figure Lengend Snippet: Diagnostic accuracy parameters of both Ag-RDTs for different definitions of RT-PCR test positivity based on viral load cut-offs, where a positive RT-PCR test with a viral load below the viral load cut-off threshold is considered a negative RT-PCR test result. Points highlighted in red indicate a viral load cut-off of 5.2 log10 E gene copies/mL, which was considered the viral load cut-off for infectiousness as determined by viral culture. BD = BD Veritor™ System for Rapid Detection of SARS-CoV-2 Ag-RDT; SD-B = Roche/SD Biosensor SARS-CoV-2 Rapid Antigen Test; PPV = positive predictive value; NPV = negative predictive value.

    Article Snippet: At the Erasmus MC Viroscience diagnostic laboratory, samples of participants with a positive RT-PCR test result were cultured for seven days, and, once cytopathic effects (CPE) were visible, the presence of SARS-CoV-2 was confirmed with immunofluorescent detection of SARS-CoV-2 nucleocapsid protein (Rabbit polyclonal antibody, Sino Biological inc.), Eschborn, Germany) (Supplementary Material 2).

    Techniques: Diagnostic Assay, Reverse Transcription Polymerase Chain Reaction

    Flow of study participants. BD = BD Veritor™ System for Rapid Detection of SARS-CoV-2 Ag-RDT (‘BD’), SD-B = Roche/SD Biosensor SARS-CoV-2 Rapid Antigen Test.

    Journal: medRxiv

    Article Title: Diagnostic accuracy of rapid antigen tests in pre-/asymptomatic close contacts of individuals with a confirmed SARS-CoV-2 infection

    doi: 10.1101/2021.03.18.21253874

    Figure Lengend Snippet: Flow of study participants. BD = BD Veritor™ System for Rapid Detection of SARS-CoV-2 Ag-RDT (‘BD’), SD-B = Roche/SD Biosensor SARS-CoV-2 Rapid Antigen Test.

    Article Snippet: At the Erasmus MC Viroscience diagnostic laboratory, samples of participants with a positive RT-PCR test result were cultured for seven days, and, once cytopathic effects (CPE) were visible, the presence of SARS-CoV-2 was confirmed with immunofluorescent detection of SARS-CoV-2 nucleocapsid protein (Rabbit polyclonal antibody, Sino Biological inc.), Eschborn, Germany) (Supplementary Material 2).

    Techniques:

    Distribution of viral loads of individuals with a positive RT-PCR test result, stratified by a combination of the Roche/SD Biosensor SARS-CoV-2 Rapid Antigen Test result (Ag-RDT+/-) and the ability to culture (culture +/-), with Ag-RDT+ and Ag-RDT-corresponding to a positive and negative Ag-RDT result, and culture+ and culture-indicating whether it was possible to culture virus or not.

    Journal: medRxiv

    Article Title: Diagnostic accuracy of rapid antigen tests in pre-/asymptomatic close contacts of individuals with a confirmed SARS-CoV-2 infection

    doi: 10.1101/2021.03.18.21253874

    Figure Lengend Snippet: Distribution of viral loads of individuals with a positive RT-PCR test result, stratified by a combination of the Roche/SD Biosensor SARS-CoV-2 Rapid Antigen Test result (Ag-RDT+/-) and the ability to culture (culture +/-), with Ag-RDT+ and Ag-RDT-corresponding to a positive and negative Ag-RDT result, and culture+ and culture-indicating whether it was possible to culture virus or not.

    Article Snippet: At the Erasmus MC Viroscience diagnostic laboratory, samples of participants with a positive RT-PCR test result were cultured for seven days, and, once cytopathic effects (CPE) were visible, the presence of SARS-CoV-2 was confirmed with immunofluorescent detection of SARS-CoV-2 nucleocapsid protein (Rabbit polyclonal antibody, Sino Biological inc.), Eschborn, Germany) (Supplementary Material 2).

    Techniques: Reverse Transcription Polymerase Chain Reaction

    DENV NS1 binds to platelets and induces activation through TLR4 signal transduction. (A) The protein expression levels of TLR4 and β actin, an internal control, in human-isolated platelets from 3 different donors were detected using Western blotting (50 μg protein/lane). (B) Human-isolated platelets were preincubated with αTLR4 or a control Rabbit IgG for 1 h, and the binding of NS1 on platelet surfaces was determined by flow cytometry using FITC-conjugated anti-NS1 monoclonal antibodies (33D2-FITC) (n = 3). (C) The binding of NS1 on platelets isolated from wild-type or TLR4 knockout mice was determined by flow cytometry (n = 4). Platelets were preincubated with or without different concentrations of (D) αTLR4 (E) the TLR4 antagonist LPS-Rs, (F) the TLR4 signaling inhibitor TAK242, or the LPS inhibitor polymyxin B (10 μg/ml) for 30 min, followed by BSA or NS1 (10 μg/ml) stimulation for 1 h (n = 4). The percent fluorescence of NS1 binding and the P-selectin surface expression on platelets were analyzed by FACSCalibur flow cytometry. *P

    Journal: PLoS Pathogens

    Article Title: Dengue virus nonstructural protein 1 activates platelets via Toll-like receptor 4, leading to thrombocytopenia and hemorrhage

    doi: 10.1371/journal.ppat.1007625

    Figure Lengend Snippet: DENV NS1 binds to platelets and induces activation through TLR4 signal transduction. (A) The protein expression levels of TLR4 and β actin, an internal control, in human-isolated platelets from 3 different donors were detected using Western blotting (50 μg protein/lane). (B) Human-isolated platelets were preincubated with αTLR4 or a control Rabbit IgG for 1 h, and the binding of NS1 on platelet surfaces was determined by flow cytometry using FITC-conjugated anti-NS1 monoclonal antibodies (33D2-FITC) (n = 3). (C) The binding of NS1 on platelets isolated from wild-type or TLR4 knockout mice was determined by flow cytometry (n = 4). Platelets were preincubated with or without different concentrations of (D) αTLR4 (E) the TLR4 antagonist LPS-Rs, (F) the TLR4 signaling inhibitor TAK242, or the LPS inhibitor polymyxin B (10 μg/ml) for 30 min, followed by BSA or NS1 (10 μg/ml) stimulation for 1 h (n = 4). The percent fluorescence of NS1 binding and the P-selectin surface expression on platelets were analyzed by FACSCalibur flow cytometry. *P

    Article Snippet: Enzyme-linked immunosorbent assay (ELISA) To examine the direct interaction between DENV NS1 and candidate receptors, 50 μl of TLR4 (purified from baculovirus-system, Sino Biological), TLR2 (purified from baculovirus-system, Sino Biological), His-taq protein or BSA (5 μg/ml) in PBS (pH 7.3) was coated onto 96-well ELISA plates at 4 °C overnight.

    Techniques: Activation Assay, Transduction, Expressing, Isolation, Western Blot, Binding Assay, Flow Cytometry, Knock-Out, Mouse Assay, Fluorescence

    TLR4 is involved in DENV-induced prolongation of bleeding time and hemorrhage in mice. A hemorrhagic mouse model was performed with C57BL/6J mice (WT) and TLR4 -/- C57BL/6J background mice as described in the Methods. (A) Mouse skin samples were removed to observe local hemorrhage on day 3 after DENV injection. The number of mice with hemorrhage divided by the total number of mice inoculated in each group is indicated. Yellow arrows indicate local skin hemorrhage. (B) The clinical score of hemorrhage was quantified and determined as digital hemorrhage severity. (C) The tail bleeding time and (D) platelet counts were also determined on day 3 before sacrifice. *P

    Journal: PLoS Pathogens

    Article Title: Dengue virus nonstructural protein 1 activates platelets via Toll-like receptor 4, leading to thrombocytopenia and hemorrhage

    doi: 10.1371/journal.ppat.1007625

    Figure Lengend Snippet: TLR4 is involved in DENV-induced prolongation of bleeding time and hemorrhage in mice. A hemorrhagic mouse model was performed with C57BL/6J mice (WT) and TLR4 -/- C57BL/6J background mice as described in the Methods. (A) Mouse skin samples were removed to observe local hemorrhage on day 3 after DENV injection. The number of mice with hemorrhage divided by the total number of mice inoculated in each group is indicated. Yellow arrows indicate local skin hemorrhage. (B) The clinical score of hemorrhage was quantified and determined as digital hemorrhage severity. (C) The tail bleeding time and (D) platelet counts were also determined on day 3 before sacrifice. *P

    Article Snippet: Enzyme-linked immunosorbent assay (ELISA) To examine the direct interaction between DENV NS1 and candidate receptors, 50 μl of TLR4 (purified from baculovirus-system, Sino Biological), TLR2 (purified from baculovirus-system, Sino Biological), His-taq protein or BSA (5 μg/ml) in PBS (pH 7.3) was coated onto 96-well ELISA plates at 4 °C overnight.

    Techniques: Mouse Assay, Injection

    Proposed mechanisms of the contribution of DENV NS1 to cause thrombocytopenia and hemorrhage during DENV infection. Circulating DENV NS1 binds to platelets via TLR4 or other molecules to induce the release of ADP, which in turn elevates P-selectin expression and PS exposure on platelet surfaces leading to platelet activation and enhancement of the platelet aggregation. In addition, NS1-activated platelets are prone to adhere onto endothelium or phagocytosis by macrophages. On the other hand, NS1 can also bind to endothelial cells and macrophage to cause their activation and cytokine release. All these effects induced by NS1 can contribute to the thrombocytopenia and hemorrhage during DENV infection.

    Journal: PLoS Pathogens

    Article Title: Dengue virus nonstructural protein 1 activates platelets via Toll-like receptor 4, leading to thrombocytopenia and hemorrhage

    doi: 10.1371/journal.ppat.1007625

    Figure Lengend Snippet: Proposed mechanisms of the contribution of DENV NS1 to cause thrombocytopenia and hemorrhage during DENV infection. Circulating DENV NS1 binds to platelets via TLR4 or other molecules to induce the release of ADP, which in turn elevates P-selectin expression and PS exposure on platelet surfaces leading to platelet activation and enhancement of the platelet aggregation. In addition, NS1-activated platelets are prone to adhere onto endothelium or phagocytosis by macrophages. On the other hand, NS1 can also bind to endothelial cells and macrophage to cause their activation and cytokine release. All these effects induced by NS1 can contribute to the thrombocytopenia and hemorrhage during DENV infection.

    Article Snippet: Enzyme-linked immunosorbent assay (ELISA) To examine the direct interaction between DENV NS1 and candidate receptors, 50 μl of TLR4 (purified from baculovirus-system, Sino Biological), TLR2 (purified from baculovirus-system, Sino Biological), His-taq protein or BSA (5 μg/ml) in PBS (pH 7.3) was coated onto 96-well ELISA plates at 4 °C overnight.

    Techniques: Infection, Expressing, Activation Assay