Journal: bioRxiv

Article Title: SARS-CoV-2 Spike triggers barrier dysfunction and vascular leak via integrins and TGF-β signaling

doi: 10.1101/2021.12.10.472112

Figure Lengend Snippet: (A) Schematic depicting S-triggered barrier dysfunction measured by a trans-endothelial/epithelial electrical resistance assay (TEER; left) and an endo/epithelial glycocalyx layer (EGL) assay (right). (B) Time course TEER assay measuring the barrier function of HPMEC monolayers over time with the indicated treatments, including DENV2 NS1 (5 μg/mL), VEGF (50 ng/mL), and SARS-CoV-2 S (10 μg/mL). Data are from n=2 biological replicates. (C) A TEER assay measuring the barrier of monolayers of HPMEC and HPMEC/ACE2 at 24 hours after the indicated treatments. VEGF positive control (50 ng/mL). Data are from n=3 biological replicates. (D) Same as C but treated with the indicated VSV pseudotyped particles at the indicated dilutions. VSV-bald and VSV-G are diluted 1:30. Data are from n=3 biological replicates. (E) Same as C but treated with the indicated concentrations of SARS-CoV-2 RBD. Data are from n=3 biological replicates. (F) Same as C but measuring the barrier of Calu-3 cell monolayers. Data are from n=2 biological replicates. (G) A TEER inhibition assay measuring the capacity of a cocktail of anti-S antibodies to inhibit S-mediated endothelial hyperpermeability. S (10 μg/mL) and the antibody cocktail (15 μg/mL for each antibody; 1A9 [Genetex] and CR3022 [Absolute Antibody]) were added simultaneously to the upper chamber of transwell inserts to a monolayer of HPMEC or HPMEC/ACE2 and TEER was measured 24 hours post-treatment (hpt). Data are from n=2 biological replicates. In all panels, the dotted line is the normalized TEER value of the untreated control condition. All data are plotted as mean +/− SD. For all panels, values are compared to untreated controls by ANOVA with multiple comparisons with *p<0.05, **p<0.01, ***p<0.001, and n.s. p>0.05.

Article Snippet: Purified proteins were formulated at ~1 mg/mL in PBS and stored in aliquots at −80° C. Recombinant DENV serotype 2 NS1 was purchased from the Native Antigen Company and characterized previously (Dengue virus serotype 2 NS1 [accession # P29990.1, Thailand/16681/84]) ( ).

Techniques: Positive Control, Inhibition

Journal: bioRxiv

Article Title: SARS-CoV-2 Spike triggers barrier dysfunction and vascular leak via integrins and TGF-β signaling

doi: 10.1101/2021.12.10.472112

Figure Lengend Snippet: (A) A representative mouse back from a dermal leak experiment. The dorsal dermises of mice were injected intradermally with the treatments and doses indicated. Mice then received a dextran-680 tracer molecule intravenously. Mouse dermises were collected 2 h post-treatment and quantification of local dermal leak was assessed by a fluorescent scanner. (B) Quantification of A from mice treated with PBS (n=27), DENV2 NS1 (15 μg; n=5), S (10 μg; n=25), and S (25 μg; n=5). (C) Representative lung images from a SARS-CoV-2 S systemic vascular leak assay. Mice were administered 50 μg of SARS-CoV-2 S or ovalbumin intranasally as indicated, and 22 hpt were administered a dextran-680 tracer intravenously as in A. Organs of mice were collected 2 hours post dextran-680 administration (24 hours post-S treatment), and accumulation of dextran-680 was measured with a fluorescent scanner. (D) Quantification of C from n=6 mice. (E) Same as C except representative images of spleens. (F) Quantification of E from n=6 mice. (G) Same as C except representative images of small intestine. (H) Quantification of G from n=5 mice. MFI is mean fluorescence intensity. All data are plotted as mean +/− SEM with *p<0.05, **p<0.01, and ***p<0.001 by unpaired t-test.

Article Snippet: Purified proteins were formulated at ~1 mg/mL in PBS and stored in aliquots at −80° C. Recombinant DENV serotype 2 NS1 was purchased from the Native Antigen Company and characterized previously (Dengue virus serotype 2 NS1 [accession # P29990.1, Thailand/16681/84]) ( ).

Techniques: Injection, Fluorescence

Journal: PLoS Neglected Tropical Diseases

Article Title: The Diagnostic Sensitivity of Dengue Rapid Test Assays Is Significantly Enhanced by Using a Combined Antigen and Antibody Testing Approach

doi: 10.1371/journal.pntd.0001199

Figure Lengend Snippet: Increasing concentrations of dengue-1 NS1 were sequentially injected onto 70 RU of Ab3 captured by anti-mouse IgG. The reference-subtracted data (black curve) were fitted using 1∶1 binding algorithm (red curve).

Article Snippet: Recombinant DENV2 NS1 was supplied by Hawaii Biotech (U.S.A).

Techniques: Injection, Binding Assay

Journal: PLoS Neglected Tropical Diseases

Article Title: The Diagnostic Sensitivity of Dengue Rapid Test Assays Is Significantly Enhanced by Using a Combined Antigen and Antibody Testing Approach

doi: 10.1371/journal.pntd.0001199

Figure Lengend Snippet: Binding kinetics of anti-NS1 MAbs to dengue NS1 antigens determined by 1∶1 binding model.

Article Snippet: Recombinant DENV2 NS1 was supplied by Hawaii Biotech (U.S.A).

Techniques: Binding Assay

Journal: PLoS Neglected Tropical Diseases

Article Title: The Diagnostic Sensitivity of Dengue Rapid Test Assays Is Significantly Enhanced by Using a Combined Antigen and Antibody Testing Approach

doi: 10.1371/journal.pntd.0001199

Figure Lengend Snippet: The test line signal intensity was measured against a serial two-fold dilution series of DENV-1 NS1 (A), DENV-2 NS1 (B), DENV-3 NS1 (C) and DENV-4 NS1 (D) from 512 ng/mL to 0.5 ng/mL. The analytical limit of detection was determined as the lowest concentration required to produce a positive result of 8.5 mABS (absorbance) units using interpolation from a non-linear regression. An absorbance value of 8.5 mABS was determined to be the cut-off value for visualisation of the test line.

Article Snippet: Recombinant DENV2 NS1 was supplied by Hawaii Biotech (U.S.A).

Techniques: Concentration Assay

Journal: PLoS Neglected Tropical Diseases

Article Title: The Diagnostic Sensitivity of Dengue Rapid Test Assays Is Significantly Enhanced by Using a Combined Antigen and Antibody Testing Approach

doi: 10.1371/journal.pntd.0001199

Figure Lengend Snippet: Accuracy of Dengue Early Rapid (NS1) test in Vietnamese study site.

Article Snippet: Recombinant DENV2 NS1 was supplied by Hawaii Biotech (U.S.A).

Techniques:

Journal: PLoS ONE

Article Title: Development of peptide biosensor for the detection of dengue fever biomarker, nonstructural 1

doi: 10.1371/journal.pone.0222144

Figure Lengend Snippet: The functionalized gold surface was exposed to a solution of synthetic peptides to create the affinity peptide-decorated electrochemical sensor for the detection of the dengue virus antigen, nonstructural 1 (NS1).

Article Snippet: Recombinant DENV NS1 proteins (DENV-1, DENV-2, DENV-3 and DENV-4, >95% purity) was purchased from Abcam (Cambridge, UK).

Techniques:

Journal: PLoS ONE

Article Title: Development of peptide biosensor for the detection of dengue fever biomarker, nonstructural 1

doi: 10.1371/journal.pone.0222144

Figure Lengend Snippet: (A) Effect of peptide concentrations. The current change at varying concentrations of DGV BP peptides (from 1 to 100 μg/mL) on a gold electrode layer was measured before and after the addition of pure dengue virus (DENV) type 2 nonstructural 1 (NS1) proteins (denoted DENV2 NS1, 12.5 μg/mL). (B) Relative current change on the binding interaction. (C) Linear correlation of the sensor with different NS1 protein concentrations, measured by square wave voltammetry. Inset shows relationship between relative I (%) with lower concentration of NS1 ranging from 0 to 1.4 μg/mL. (D) Stability test of affinity peptide-decorated sensor.

Article Snippet: Recombinant DENV NS1 proteins (DENV-1, DENV-2, DENV-3 and DENV-4, >95% purity) was purchased from Abcam (Cambridge, UK).

Techniques: Binding Assay, Concentration Assay

Journal: PLoS ONE

Article Title: Development of peptide biosensor for the detection of dengue fever biomarker, nonstructural 1

doi: 10.1371/journal.pone.0222144

Figure Lengend Snippet: Current change was measured by square wave voltammetry before and after the addition of four pure DENV nonstructural 1 (NS1) proteins (12.5 μg/mL) and culture broth from DENV serotypes 1–4 to a DGV BP1 peptide-decorated gold electrode layer.

Article Snippet: Recombinant DENV NS1 proteins (DENV-1, DENV-2, DENV-3 and DENV-4, >95% purity) was purchased from Abcam (Cambridge, UK).

Techniques:

Journal: Cell reports

Article Title: Antigenic Variation of the Dengue Virus 2 Genotypes Impacts the Neutralization Activity of Human Antibodies in Vaccinees

doi: 10.1016/j.celrep.2020.108226

Figure Lengend Snippet: (A) Phylogenetic relationships of the prM and E sequences among the six dengue virus serotype 2 genotypes: American, Asian I, Asian II, Asian-American, Cosmopolitan, and Sylvatic. Colored circles represent DENV2 genotypic isolates that were produced recombinantly. Red arrow represents the dengue virus 2 NIH vaccine strain, and the blue arrow shows the DENV2 Dengvaxia vaccine strain. (B) Schematic representation of recombinant DENV2 prM and E viral constructs on the S16803 WHO reference strain backbone. See also .

Article Snippet: Recombinant envelope protein (recE) (80% of E protein) from WHO reference DENV2 S16803 strain was produced in our laboratory or purchased from Hawaii Biotech, Inc.( ) Recombinant EDIII protein from DENV2 was produced in-house as described previously ( ).

Techniques: Virus, Produced, Recombinant, Construct

Journal: Cell reports

Article Title: Antigenic Variation of the Dengue Virus 2 Genotypes Impacts the Neutralization Activity of Human Antibodies in Vaccinees

doi: 10.1016/j.celrep.2020.108226

Figure Lengend Snippet: (A) DENV2 genotype variant foci size in Vero-81 cells at 48 h post-infection. (B) Quantification of foci size (1e −3 mm 2 ). (C) Multistep growth curve kinetics of dengue virus 2 genotypic variants in C6/36 insect cells. Cells were infected with a multiplicity of infection of 0.01. p values displayed are from a Kruskal-Wallis test. (D) Stability of the DENV2 genotype panel at 4°C, 28°C, 37°C, and 40°C. Mean percentage of infection is shown from two independent repeats. Bars denote the range. See also and .

Article Snippet: Recombinant envelope protein (recE) (80% of E protein) from WHO reference DENV2 S16803 strain was produced in our laboratory or purchased from Hawaii Biotech, Inc.( ) Recombinant EDIII protein from DENV2 was produced in-house as described previously ( ).

Techniques: Variant Assay, Infection, Virus

Journal: Cell reports

Article Title: Antigenic Variation of the Dengue Virus 2 Genotypes Impacts the Neutralization Activity of Human Antibodies in Vaccinees

doi: 10.1016/j.celrep.2020.108226

Figure Lengend Snippet: (A) Amino acid variation within the prM. (B) Amino acid variation within the EDI, EDII, and EDIII among distinct DENV2 genotypes from different parts of the world. (C) Amino acid residue variation on the E protein surface. Amino acid residue variation hotspots relative to S16803 WHO reference strain. See also .

Article Snippet: Recombinant envelope protein (recE) (80% of E protein) from WHO reference DENV2 S16803 strain was produced in our laboratory or purchased from Hawaii Biotech, Inc.( ) Recombinant EDIII protein from DENV2 was produced in-house as described previously ( ).

Techniques: Residue

Journal: Cell reports

Article Title: Antigenic Variation of the Dengue Virus 2 Genotypes Impacts the Neutralization Activity of Human Antibodies in Vaccinees

doi: 10.1016/j.celrep.2020.108226

Figure Lengend Snippet: (A and B) Memory-B-cell-derived mAbs 2D22 and 3F9. (C–H) Plasmablast-derived mAbs 1662, 1671-1, 1671-4, 1615, 1628, and 1660. (I) mAb neutralization IC 50 values against DENV2 recombinant viruses. See also .

Article Snippet: Recombinant envelope protein (recE) (80% of E protein) from WHO reference DENV2 S16803 strain was produced in our laboratory or purchased from Hawaii Biotech, Inc.( ) Recombinant EDIII protein from DENV2 was produced in-house as described previously ( ).

Techniques: Derivative Assay, Neutralization, Recombinant

Journal: Cell reports

Article Title: Antigenic Variation of the Dengue Virus 2 Genotypes Impacts the Neutralization Activity of Human Antibodies in Vaccinees

doi: 10.1016/j.celrep.2020.108226

Figure Lengend Snippet: (A) The neutralization activity in individuals immunized with the TV003 NIH DENV2 monovalent vaccine. Horizontal dashed line denotes the starting dilution tested. (B) The neutralization activity in individuals immunized with theTV003 NIH DENV tetravalent vaccine against DENV2 viruses. p values are from a Friedman’s test. See also and and .

Article Snippet: Recombinant envelope protein (recE) (80% of E protein) from WHO reference DENV2 S16803 strain was produced in our laboratory or purchased from Hawaii Biotech, Inc.( ) Recombinant EDIII protein from DENV2 was produced in-house as described previously ( ).

Techniques: Neutralization, Activity Assay

Journal: Cell reports

Article Title: Antigenic Variation of the Dengue Virus 2 Genotypes Impacts the Neutralization Activity of Human Antibodies in Vaccinees

doi: 10.1016/j.celrep.2020.108226

Figure Lengend Snippet: (A) The neutralization activity of BSA-depleted DENV2-specific IgG neutralizing antibodies from experimentally infected individuals. (B) The neutralization activity of cross-reactive antibody-depleted DENV2-specific IgG neutralizing antibodies in experimentally infected individuals. (C) Percent DENV2-serotype-specific neutralizing antibodies in experimentally infected individuals following cross-reactive antibody depletion. Bars denote mean and standard deviation. See also .

Article Snippet: Recombinant envelope protein (recE) (80% of E protein) from WHO reference DENV2 S16803 strain was produced in our laboratory or purchased from Hawaii Biotech, Inc.( ) Recombinant EDIII protein from DENV2 was produced in-house as described previously ( ).

Techniques: Neutralization, Activity Assay, Infection, Standard Deviation

Journal: Cell reports

Article Title: Antigenic Variation of the Dengue Virus 2 Genotypes Impacts the Neutralization Activity of Human Antibodies in Vaccinees

doi: 10.1016/j.celrep.2020.108226

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Recombinant envelope protein (recE) (80% of E protein) from WHO reference DENV2 S16803 strain was produced in our laboratory or purchased from Hawaii Biotech, Inc.( ) Recombinant EDIII protein from DENV2 was produced in-house as described previously ( ).

Techniques: Virus, Recombinant, Saline, Modification, Software