Journal: bioRxiv

Article Title: Capturing and Recreating Diverse Antibody Repertoires as Multivalent Recombinant Polyclonal Antibody Drugs

doi: 10.1101/2020.08.05.232975

Figure Lengend Snippet: ( a ) Clonal cluster analysis of rZIG-IgG1 (blue) and rZIG-LALA (green) antibodies. Each node represents an antibody clone (full-length heavy chain). The size of the nodes reflects the frequency of the scFv clones in the final library (only clones ≥0.01% are plotted). We computed the total number of amino acid differences between each pairwise alignment after combining both libraries together, and edges indicate ≤5 amino acid differences. ( b ) ELISA of rZIG-IgG1 (blue), rZIG-LALA (green), and Zika/Dengue+ serum control (red) for Dengue serotypes 1-4 (y-axis; indicated by shape) and Zika virus antigen (x-axis). Linear regression trendline is indicated in black. ( c ) Pseudotype neutralization by rZIG-IgG1 (blue), rZIG-LALA (green), and Zika/Dengue+ serum control (red) for Dengue serotypes 1-4 (y-axis; indicated by shape) and Zika virus antigen (x-axis). Linear regression trendline is indicated in black. ( d ) Zika pseudotype virus ADE assay for rZIG-IgG1 (blue), rZIG-LALA (green), and positive and negative controls. Test article concentration is on the x-axis. Fold-increase infection is on the y-axis, which was the infection-induced luciferase signal observed in the presence of antibody divided by the luciferase signal observed with a no antibody control.

Article Snippet: After blocking the coated plate with ultrablock buffer (Bio-Rad, Hercules, CA, USA) and washing with PBS + 0.05% Tween-20 (Teknova, Hollister, CA, USA), eight-step three-fold serial dilutions in assay buffer (1 × PBS + 0.05% Tween + 0.3% dry milk) were performed on rZIG-IgG1, rZIG-LALA, Zika/Dengue+ serum positive control (Seracare, Milford, MA, USA), and a negative control IVIG (Gamunex; Grifols, S.A., Sant Cugat, Spain).

Techniques: Clone Assay, Enzyme-linked Immunosorbent Assay, Neutralization, Concentration Assay, Infection, Luciferase

Journal: bioRxiv

Article Title: Capturing and Recreating Diverse Antibody Repertoires as Multivalent Recombinant Polyclonal Antibody Drugs

doi: 10.1101/2020.08.05.232975

Figure Lengend Snippet: rZIG-IgG1 (blue), rZIG-LALA (green), negative control IVIG (black), and positive control Zika/Dengue positive serum (red) were serially diluted and added to a Zika envelope-coated plate. Antigen-specific responses were quantified by an anti-human-HRP secondary antibody.

Article Snippet: After blocking the coated plate with ultrablock buffer (Bio-Rad, Hercules, CA, USA) and washing with PBS + 0.05% Tween-20 (Teknova, Hollister, CA, USA), eight-step three-fold serial dilutions in assay buffer (1 × PBS + 0.05% Tween + 0.3% dry milk) were performed on rZIG-IgG1, rZIG-LALA, Zika/Dengue+ serum positive control (Seracare, Milford, MA, USA), and a negative control IVIG (Gamunex; Grifols, S.A., Sant Cugat, Spain).

Techniques: Negative Control, Positive Control

Journal: bioRxiv

Article Title: Capturing and Recreating Diverse Antibody Repertoires as Multivalent Recombinant Polyclonal Antibody Drugs

doi: 10.1101/2020.08.05.232975

Figure Lengend Snippet: rZIG-IgG1 (blue), rZIG-LALA (green), negative control IVIG (black), and positive control Zika/Dengue positive serum (red) were serially diluted and added to a Dengue serotypes 1, 2, 3, or 4 envelope-coated plates. Antigen-specific responses were quantified by an anti-human-HRP secondary antibody.

Article Snippet: After blocking the coated plate with ultrablock buffer (Bio-Rad, Hercules, CA, USA) and washing with PBS + 0.05% Tween-20 (Teknova, Hollister, CA, USA), eight-step three-fold serial dilutions in assay buffer (1 × PBS + 0.05% Tween + 0.3% dry milk) were performed on rZIG-IgG1, rZIG-LALA, Zika/Dengue+ serum positive control (Seracare, Milford, MA, USA), and a negative control IVIG (Gamunex; Grifols, S.A., Sant Cugat, Spain).

Techniques: Negative Control, Positive Control

Journal: bioRxiv

Article Title: Capturing and Recreating Diverse Antibody Repertoires as Multivalent Recombinant Polyclonal Antibody Drugs

doi: 10.1101/2020.08.05.232975

Figure Lengend Snippet: rZIG-IgG1 (blue), rZIG-LALA (green), negative control IVIG (black), positive control Zika/Dengue mAb (gray), and positive control Zika/Dengue positive serum (red) were serially diluted, co-incubated with Zika pseudotype virus, and added to BHK/DC-SIGN target cells. Antibody induced neutralization was quantified by the infection-induced luciferase expression divided by luciferase expression in the no Ab control.

Article Snippet: After blocking the coated plate with ultrablock buffer (Bio-Rad, Hercules, CA, USA) and washing with PBS + 0.05% Tween-20 (Teknova, Hollister, CA, USA), eight-step three-fold serial dilutions in assay buffer (1 × PBS + 0.05% Tween + 0.3% dry milk) were performed on rZIG-IgG1, rZIG-LALA, Zika/Dengue+ serum positive control (Seracare, Milford, MA, USA), and a negative control IVIG (Gamunex; Grifols, S.A., Sant Cugat, Spain).

Techniques: Negative Control, Positive Control, Incubation, Neutralization, Infection, Luciferase, Expressing

Journal: bioRxiv

Article Title: Capturing and Recreating Diverse Antibody Repertoires as Multivalent Recombinant Polyclonal Antibody Drugs

doi: 10.1101/2020.08.05.232975

Figure Lengend Snippet: rZIG-IgG1 (blue), rZIG-LALA (green), negative control IVIG (black), positive control Zika/Dengue mAb (gray), and positive control Zika/Dengue positive serum (red) were serially diluted, co-incubated with pseudotype virus expressing the indicated Dengue serotype envelope antigens, and added to BHK/DC-SIGN target cells. Antibody induced neutralization was quantified as “% no Ab control”, or infection-induced luciferase expression divided by the luciferase expression in the no Ab control.

Article Snippet: After blocking the coated plate with ultrablock buffer (Bio-Rad, Hercules, CA, USA) and washing with PBS + 0.05% Tween-20 (Teknova, Hollister, CA, USA), eight-step three-fold serial dilutions in assay buffer (1 × PBS + 0.05% Tween + 0.3% dry milk) were performed on rZIG-IgG1, rZIG-LALA, Zika/Dengue+ serum positive control (Seracare, Milford, MA, USA), and a negative control IVIG (Gamunex; Grifols, S.A., Sant Cugat, Spain).

Techniques: Negative Control, Positive Control, Incubation, Expressing, Neutralization, Infection, Luciferase