Journal: Immunology

Article Title: Non-structural protein 1-specific antibodies directed against Zika virus in humans mediate antibody-dependent cellular cytotoxicity.

doi: 10.1111/imm.13380

Figure Lengend Snippet: FIGURE 1 Isotype specificity of ZIKV-immune sera. ELISA plates were coated with ZIKV NS1 (a, c) or ZIKV VLP (b, d), and a 1:100 dilution (n = 28) (a, b) or serial dilutions (n = 11) (c, d) of ZIKV-immune sera were added to plates. Following incubation and washes, optimized concentrations of polyclonal secondary IgG, and monoclonal Abs against the hinge or Fc portion of IgG1, IgG2, IgG3 and IgG4 Abs were added to wells (a) and (b) and IgG1 hinge Abs to (c) and (d). OD values shown at 450 nm. OD values to ZIKV VLP and ZIKV NS1 using a 1:100 dilution of ZIKV-naïve sera (n = 4), <0·2 with polyclonal secondary IgG and <0·1 with IgG1 hinge, IgG1 Fc, IgG2, IgG3 and IgG4 secondary Abs. Bars represent mean with standard error of the mean of the OD values. *P < 0·05, ***P < 0·001 and ****P < 0·0001 were obtained using the non- parametric Friedman test (three or more matched groups) with Dunn's multiple comparisons test

Article Snippet: Briefly, 96- well plates were coated overnight with 20 ng/well of ZIKV virus- like particles (Suriname Z1106033) or 50 ng/ml ZIKV NS1 (Uganda MR766) (Native Antigen Company, Oxfordshire, UK).

Techniques: Enzyme-linked Immunosorbent Assay, Incubation

Journal: Immunology

Article Title: Non-structural protein 1-specific antibodies directed against Zika virus in humans mediate antibody-dependent cellular cytotoxicity.

doi: 10.1111/imm.13380

Figure Lengend Snippet: FIGURE 2 Frequencies of ZIKV-specific IgG and IgG1 B-cell responses using fluorescently labelled ZIKV or ZIKV NS1. PBMCs from ZIKV-immune and ZIKV-naïve donors were stimulated in vitro for seven days with r848 + rIL-2. FluoroSpot plates were coated with IgG and IgG1 capture Abs. (a) Images of wells containing media, total IgG, ZIKV and ZIKV NS1 IgG- or IgG1-specific MBCs from one representative donor. (b) Frequencies of ZIKV (green) and NS1-specific (red) ASCs per 106 input cells in MBC cultures from 12 ZIKV-immune and three ZIKV- naïve donors. Values are the mean of duplicate wells for each condition. The supernatants were evaluated for isotype-specific Abs to (c) ZIKV VLPs and (D) ZIKV NS1. *P < 0·05, **P < 0·01 and ****P < 0·0001 were obtained using the non-parametric Mann-Whitney or Friedman test (three or more matched groups) with Dunn's multiple comparisons test

Article Snippet: Briefly, 96- well plates were coated overnight with 20 ng/well of ZIKV virus- like particles (Suriname Z1106033) or 50 ng/ml ZIKV NS1 (Uganda MR766) (Native Antigen Company, Oxfordshire, UK).

Techniques: In Vitro, MANN-WHITNEY

Journal: Immunology

Article Title: Non-structural protein 1-specific antibodies directed against Zika virus in humans mediate antibody-dependent cellular cytotoxicity.

doi: 10.1111/imm.13380

Figure Lengend Snippet: FIGURE 3 Opsonization and ADCC activity of a ZIKV NS1 monoclonal Ab. (a) K562 cells transfected with DC-SIGN were infected with ZIKV (moi = 0·1 and 1), and 24 h later, the expression of E and NS1 was measured with monoclonal Abs 4G2 and B4 respectively. (b) Opsonization of ZIKV NS1-transfected CEM-NKR cells with a mAb to ZIKV NS1 from Native Antigen Company. CEM-NKR cells transfected with ZIKV NS1 (c and d) or control CEM-NKR cells (e and f) were opsonized with a ZIKV NS1 mAb (d and f), added to PBMC. A standard lymphocyte gate based upon light scatter properties followed by selection of singlet viable cells and dump channel exclusion (CD3− T cells) defined the non-T-cell lymphocyte population, and Abs to CD16 and CD56 identified NK cells

Article Snippet: Briefly, 96- well plates were coated overnight with 20 ng/well of ZIKV virus- like particles (Suriname Z1106033) or 50 ng/ml ZIKV NS1 (Uganda MR766) (Native Antigen Company, Oxfordshire, UK).

Techniques: Activity Assay, Transfection, Infection, Expressing, Control, Selection

Journal: Immunology

Article Title: Non-structural protein 1-specific antibodies directed against Zika virus in humans mediate antibody-dependent cellular cytotoxicity.

doi: 10.1111/imm.13380

Figure Lengend Snippet: FIGURE 4 NK cell activation and degranulation in the presence of ZIKV- immune sera. 1:100 dilution and 1:1600 dilution of polyclonal sera from ZIKV- immune individuals were evaluated for their ability to mediate ADCC of ZIKV NS1 CEM-NKR cells using intracellular cytokine assays. Shown are the frequencies of NK cells that (a) secrete IFN-γ, (B) express CD107 or (C) express both CD107 and IFN-γ in the presence of ZIKV-immune sera. Background levels were subtracted

Article Snippet: Briefly, 96- well plates were coated overnight with 20 ng/well of ZIKV virus- like particles (Suriname Z1106033) or 50 ng/ml ZIKV NS1 (Uganda MR766) (Native Antigen Company, Oxfordshire, UK).

Techniques: Activation Assay

Journal: Immunology

Article Title: Non-structural protein 1-specific antibodies directed against Zika virus in humans mediate antibody-dependent cellular cytotoxicity.

doi: 10.1111/imm.13380

Figure Lengend Snippet: FIGURE 5 Opsonization and target cell lysis using an NK-TVA image cytometry assay. (a) Opsonization of ZIKV NS1 CEM-NKR cells by ZIKV convalescent sera. (b) Representative images of wells acquired by an ImmunoSpot S5 Analyser of an NK-TVA assay where ethanol, ZIKV NS1 or IgG Ab was added to the wells containing effector and target cells. (c) Using an optimized number of target cells, effector PBMC as a source of NK cells and different dilutions of naïve sera (n = 5), positive control sera that opsonized ZIKV NS1 CEM-NKR targets (n = 5) and experimental sera (n = 12), the loss of target cells at the single-cell level was measured by the NK-TVA image cytometry assay and calculated as a percentage of lysed target cells. *P < 0·05 and **P < 0·01 were obtained using a Kruskal–Wallis test with Dunn's multiple comparisons test

Article Snippet: Briefly, 96- well plates were coated overnight with 20 ng/well of ZIKV virus- like particles (Suriname Z1106033) or 50 ng/ml ZIKV NS1 (Uganda MR766) (Native Antigen Company, Oxfordshire, UK).

Techniques: Lysis, Cytometry, Positive Control

Journal: Cell reports

Article Title: Fusion peptide priming reduces immune responses to HIV-1 envelope trimer base

doi: 10.1016/j.celrep.2021.108937

Figure Lengend Snippet:

Article Snippet: pVRC8400 vector , https://www.addgene.org , Cat# 63160.

Techniques: Neutralization, Recombinant, Transfection, Plasmid Preparation, Software