recombinant yfv ns1 protein  (Native Antigen Inc)

Journal: EBioMedicine

Article Title: Yellow fever disease severity and endothelial dysfunction are associated with elevated serum levels of viral NS1 protein and syndecan-1.

doi: 10.1016/j.ebiom.2024.105409

Figure Lengend Snippet: Fig. 1: Development of quantitative YFV NS1 ELISA. (a) Sigmoidal standard curve of YFV NS1 capture ELISA performed with in-house-produced YFJ19 mAb and thirteen different concentrations of recombinant YFV NS1. The graph depicts mean absorbance values ± standard deviation (SD). (b) Comparison of standard curve of YFV NS1 diluted or not in normal human serum (1:10). The graph depicts mean absorbance values ± SD. (c) Specificity of mAb YFJ19 by direct ELISA performed with NS1 (5 μg/mL) from 12 different flaviviruses as follows: YFV; dengue virus serotypes 1 (DENV1), 2 (DENV2), 3 (DENV2), and 4 (DENV4); Saint Louis encephalitis virus (SLEV); West Nile virus (WNV); Zika virus (ZIKV); Wesselsbron virus (WBV); Usutu virus (USUV); Japanese encephalitis virus (JEV); Tick-borne encephalitis virus (TBEV). (d) Cross-reactivity of anti- flavivirus NS1 mAb (2B7) by direct ELISA performed with NS1 (5 μg/mL) from the 12 different flaviviruses as in c. (e) Western blot analysis showing specificity of mAb YFJ19 for YFV NS1 and cross-reactivity of mAb 2B7.

Article Snippet: Recombinant YFV NS1 (Native Antigen Co.) at 10 μg/mL was used as a positive control.

Techniques: Enzyme-linked Immunosorbent Assay, Produced, Recombinant, Standard Deviation, Comparison, Direct ELISA, Virus, Western Blot

Journal: EBioMedicine

Article Title: Yellow fever disease severity and endothelial dysfunction are associated with elevated serum levels of viral NS1 protein and syndecan-1.

doi: 10.1016/j.ebiom.2024.105409

Figure Lengend Snippet: Fig. 2: YFV NS1 serum levels in severe and non-severe YF patients. (a) YFV NS1 levels were determined using an in-house sandwich ELISA, as described in Materials and Methods. Studied individuals were classified in three different groups as described in Materials and Methods: Severe YF (n = 39); Non-severe YF (n = 17, missing data for 1 sample); Controls, healthy individuals (n = 5, missing data for 6 samples). Groups were compared by Kruskal–Wallis + Dunn’s multiple comparisons test. (b) YFV NS1 levels in survivors and deceased YFV-infected groups. Groups were compared by Mann–Whitney test. The dashed lines indicate the median of the data, and the error bars represent the interquartile range (IQR) in a and b. (c) YFV NS1 serum levels in individuals with acute YF according to days since symptom onset, visualized using LOESS curves with 95% confidence intervals (shaded areas). Acute disease was defined by detection of viral RNA by RT-qPCR in serum. Samples were run in duplicate.

Article Snippet: Recombinant YFV NS1 (Native Antigen Co.) at 10 μg/mL was used as a positive control.

Techniques: Sandwich ELISA, Infection, MANN-WHITNEY, Quantitative RT-PCR

Journal: EBioMedicine

Article Title: Yellow fever disease severity and endothelial dysfunction are associated with elevated serum levels of viral NS1 protein and syndecan-1.

doi: 10.1016/j.ebiom.2024.105409

Figure Lengend Snippet: Fig. 4: Relative TEER of human endothelial cells treated with acute YF human serum samples and effects of YFV NS1 treatment on endothelial glycocalyx. (a) Confluent monolayers of human endothelial cells cultured in Transwell inserts were treated or not with 10% serum from three different groups: Severe YF (n = 24, missing data for 15 samples); Non-severe YF (n = 10, missing data for 8 samples); Controls (n = 11): healthy individuals. YFV NS1 (10 μg/mL) was used as positive control. The transendothelial electrical resistance (TEER) was measured from 2 to 10 h post-treatment. Graph shows mean ± SD of relative TEER for each treatment group. (b) Area under the curve (AUC) of negative peaks for the effects of the different treatments on TEER. AUC values represent the sum of the trapezoidal areas (taking into account only the negative peaks) under the curve formed by the data points. Median values of AUC for each group were compared using Kruskal–Wallis’ test. Error bars represent interquartile range (IQR). Outliers were defined according to Tukey’s method, which identifies outliers using the interquartile range (IQR; i.e., the difference between the first and third quartiles [Q3 - Q1]) as any data point below the first quartile (25th percentile) minus 1.5 times the IQR (Q1 - 1.5*IQR) or above the third quartile (75th percentile) plus 1.5 times the IQR (Q3 + 1.5*IQR). (c, d) Human endothelial cell monolayers grown on gelatin-coated coverslips were treated with medium only or 10 μg/mL of YFV NS1 for 3 h. After fixation, cell surface syndecan-1 was stained in red and nuclei in blue (Hoechst). (c) Representative figures of cell monolayers stained for syndecan-1 in red and nuclei in blue. (d) Quantification of syndecan-1 protein on the cells surface expressed as mean fluorescence intensity (MFI). NS1 treatment was compared to medium-only treatment by unpaired t-test. Error bars present SD of the mean. (e, f, g) Human endothelial cell monolayers grown on gelatin-coated coverslips were treated with medium only or with different con- centrations of YFV NS1 for 6 h. (e) Quantification of syndecan-1 in cell supernatants by ELISA. An in-house-produced mouse anti-YFV NS1 polyserum with or without 2.5 μg/mL of YFV NS1 was used as a control. Treatments were compared with medium-only treatment by one- way ANOVA + Dunnett’s test. Error bars present SD of the mean. (f) Representative figures of cell monolayers stained for sialic acid in red and nuclei in blue. (g) Quantification of sialic acid on the cell surface expressed as MFI. NS1 treatments were compared to medium-only treatment by one-way ANOVA + Dunnett’s multiple comparison test. Error bars present SD of the mean. Significant p-values are shown in the graphs. Acute YF disease was defined by detection of viral RNA by RT-qPCR in serum. Samples were run in duplicate for the TEER assays. IFA and ELISA data are the results of three independent experiments (n = 3).

Article Snippet: Recombinant YFV NS1 (Native Antigen Co.) at 10 μg/mL was used as a positive control.

Techniques: Cell Culture, Positive Control, Staining, Enzyme-linked Immunosorbent Assay, Produced, Control, Comparison, Quantitative RT-PCR

Journal: EBioMedicine

Article Title: Yellow fever disease severity and endothelial dysfunction are associated with elevated serum levels of viral NS1 protein and syndecan-1.

doi: 10.1016/j.ebiom.2024.105409

Figure Lengend Snippet: Fig. 5: Correlation analysis of clinical and laboratory data of study participants during acute YF and viral load according to days of symptoms. (a) Correlation matrix analyzed by Spearman rank correlation. Asterisks indicate significant correlations (p < 0.05, Spearman r > 0.35). Categorical variables (e.g., sex [male] and death) were entered as coded variables. (b) Correlation between serum levels of YFV NS1 and syndecan-1. (c) Correlation between serum levels of YFV NS1 and TEER AUC values. (d) Correlation between serum levels of syndecan-1 and TEER AUC values. The gray shaded area represents the 95% confidence interval for the fitted LOESS curve in b, c, and d. Abbreviations: TEER, transendothelial electrical resistance; AUC, area under the curve; Hb, hemoglobin; Ht, hematocrit; AST, aspartate transaminase; ALT, alanine aminotransferase; TPI/INR thrombin potential index/international normalized ratio; aPTT, activated partial thromboplastin time; TB, total bilirubin; DB, direct bilirubin; IB, indirect bilirubin. (e) Viral load of YF patients from severe and non-severe cases. Viral load, determined in YF patients serum at the time of admission by RT-qPCR, is expressed as genomic copies per mL. Results were plotted according to days since symptom onset reported at the time of hospital admission. Acute disease was defined by detection of viral RNA by RT-qPCR in serum.

Article Snippet: Recombinant YFV NS1 (Native Antigen Co.) at 10 μg/mL was used as a positive control.

Techniques: Quantitative RT-PCR