Human Anti-ZIKV Recombinant Antibody (clone SIgN-3C) (CAT#: PABS-0167)

Figure 1 Neutralization profile of SIgN-3C IgG or Fab to DENV2 and ZIKV in the pre- and post-attachment neutralization assay.

IgG SIgN-3C neutralizes DENV2 almost equally well in both assays, whereas ZIKV was only neutralized in the pre-attachment assay. When Fab was used, the Fab remained highly neutralizing to DENV2 but completely lost its neutralization capability against ZIKV.

Zhang, S., Loy, T., Ng, T. S., Lim, X. N., Chew, S. Y. V., Tan, T. Y., ... & Lok, S. M. (2020). A human antibody neutralizes different flaviviruses by using different mechanisms. Cell Reports, 31(4), 107584.

Figure 2 SIgN-3C inhibits fusion of DiD-labeled DENV2 with liposome.

At different concentrations of SIgN-3C IgG, DENV2 fusion to liposome was inhibited at pH 5.0. An isotype control IgG was included in the experiment and was used for normalization. Means and standard deviations from three individual experiments are shown.

Zhang, S., Loy, T., Ng, T. S., Lim, X. N., Chew, S. Y. V., Tan, T. Y., ... & Lok, S. M. (2020). A human antibody neutralizes different flaviviruses by using different mechanisms. Cell Reports, 31(4), 107584.

Figure 3 Quantification of DiD+ U937-DC-SIGN cells (antigen-presenting cell [APC] channel) by flow cytometry.

DiD-labeled DENV2 (DENV2-DiD) was incubated with or without SIgN-3C (100 mg/mL) at 37°C for 30 min followed by U937-DC SIGN adsorption of DENV2-DiD at 37°C for 30 min.

Zhang, S., Loy, T., Ng, T. S., Lim, X. N., Chew, S. Y. V., Tan, T. Y., ... & Lok, S. M. (2020). A human antibody neutralizes different flaviviruses by using different mechanisms. Cell Reports, 31(4), 107584.

Figure 4 SIgN-3C IgG has similar binding affinities to ZIKV and DENV2 virus particles.

The binding characteristics were measured by biolayer interferometry (BLI) on an Octet-Red (ForteBio) system. The dissociation constants (KD) of SIgN-3C IgG to DENV2 and ZIKV were 56 and 60 nM, respectively.

Zhang, S., Loy, T., Ng, T. S., Lim, X. N., Chew, S. Y. V., Tan, T. Y., ... & Lok, S. M. (2020). A human antibody neutralizes different flaviviruses by using different mechanisms. Cell Reports, 31(4), 107584.

Figure 5 SIgN-3C IgG has similar binding affinities to ZIKV and DENV2 virus particles.

Dynamic light scattering experiments measuring the fold change of hydrodynamic size of DENV2 and ZIKV when complexed with SIgN-3C IgG. ZIKV, but not DENV2, forms large aggregates when IgG:virion molar ratio is higher than 100:1.

Zhang, S., Loy, T., Ng, T. S., Lim, X. N., Chew, S. Y. V., Tan, T. Y., ... & Lok, S. M. (2020). A human antibody neutralizes different flaviviruses by using different mechanisms. Cell Reports, 31(4), 107584.

Figure 6 Neutralizing capacities of selected human DENV mAbs against ZIKV in vitro.

ZIKV was preincubated with serial dilutions of human DENV mAbs SIgN-3C prior to infecting Vero-E6 cells at MOI of 10. Mock-infected and virus-only conditions were used as controls. Infectivity was quantified 48 hours after infection by immunofluorescence. Data are presented as mean ± SEM of 3 to 4 independent experiments, normalized to virus-only control. Nonlinear regression fitting was used to determine the IC₅₀ values.

Kam, Y. W., Lee, C. Y. P., Teo, T. H., Howland, S. W., Amrun, S. N., Lum, F. M., ... & Rénia, L. (2017). Cross-reactive dengue human monoclonal antibody prevents severe pathologies and death from Zika virus infections. JCI insight, 2(8).

Figure 7 Binding curves of selected mAbs by ZIKV virion ELISA. OD values were normalized to the result at 30 μg/ml mAb.

Kam, Y. W., Lee, C. Y. P., Teo, T. H., Howland, S. W., Amrun, S. N., Lum, F. M., ... & Rénia, L. (2017). Cross-reactive dengue human monoclonal antibody prevents severe pathologies and death from Zika virus infections. JCI insight, 2(8).

Figure 8 Antibody-dependent enhancement (ADE) of Zika virus (ZIKV) infection.

ZIKV was preincubated with serial dilutions of 1B-H1L1, 2F-H1L3, SIgN-3C, and SIgN-3C-LALA (0.03 ng/ml to 30 μg/ml) before infecting K562 cells at MOI of 10. Noninfected cells and virus infection in the absence of mAb (control infection, gray dotted line) were used as controls. Results are presented as mean ± SEM of virus titer fold increase with the presence of different concentrations of mAbs, relative to control infection.

Kam, Y. W., Lee, C. Y. P., Teo, T. H., Howland, S. W., Amrun, S. N., Lum, F. M., ... & Rénia, L. (2017). Cross-reactive dengue human monoclonal antibody prevents severe pathologies and death from Zika virus infections. JCI insight, 2(8).

Figure 9 Binding curves of SIgN-3C and SIgN03C-LALA mAbs by ZIKV virion ELISA.

OD values were normalized to the result at 3 μg/ml.

Kam, Y. W., Lee, C. Y. P., Teo, T. H., Howland, S. W., Amrun, S. N., Lum, F. M., ... & Rénia, L. (2017). Cross-reactive dengue human monoclonal antibody prevents severe pathologies and death from Zika virus infections. JCI insight, 2(8).

Figure 10 SIgN-3C-LALA antibody preserves good neutralizing activity against ZIKV.

Data are presented as mean ±SEM of 4 independent experiments, normalized to virus-only control.Nonlinear regression fitting was used to determine the IC₅₀ values.

Kam, Y. W., Lee, C. Y. P., Teo, T. H., Howland, S. W., Amrun, S. N., Lum, F. M., ... & Rénia, L. (2017). Cross-reactive dengue human monoclonal antibody prevents severe pathologies and death from Zika virus infections. JCI insight, 2(8).

Figure 11 (A) Mortality, (B) weight change, and (C) viremia of Zika virus-infected (ZIKV-infected) type I interferon receptor-deficient (IFNAR^-/-) mice receiving isotype (n = 10), SIgN-3C (n = 5), and SIgN3C-LALA (n = 10) antibodies.

Mice were inoculated with 10⁴ PFU ZIKV s.c. at the ventral side of the footpad and treatments were given on days 1, 4, and 8 after infection. Mice were treated with 1 mg per dose of SIgN-3C and 0.5 mg per dose of SIgN-3C-LALA.

Kam, Y. W., Lee, C. Y. P., Teo, T. H., Howland, S. W., Amrun, S. N., Lum, F. M., ... & Rénia, L. (2017). Cross-reactive dengue human monoclonal antibody prevents severe pathologies and death from Zika virus infections. JCI insight, 2(8).

Figure 12 (D) Viral load in brain, spleen, liver, kidney, testes, and popliteal lymph node (pLN) of ZIKV-infected IFNAR^-/- mice receiving isotype (n = 5), SIgN-3C (n = 5), and SIgN-3C-LALA (n = 4) antibodies at 6 days after infection.

The mortality curve was analyzed using a log-rank (Mantel-Cox) test, while weight change, viremia, and viral load were analyzed using the Kruskal-Wallis test with Dunn's multiple comparison. ^*P < 0.05; ^**P < 0.01; ^***P < 0.001.

Kam, Y. W., Lee, C. Y. P., Teo, T. H., Howland, S. W., Amrun, S. N., Lum, F. M., ... & Rénia, L. (2017). Cross-reactive dengue human monoclonal antibody prevents severe pathologies and death from Zika virus infections. JCI insight, 2(8).

Figure 13 (A) Representative images and (B) weight of fetuses isolated from mock-infected type I interferon receptor-deficient (IFNAR^-/-) mice, Zika virus-infected (ZIKV-infected) isotype control, and ZIKV-infected mice with SIgN-3C or SIgN-3C-LALA treatments (all groups n ≥ 7).

Each data point in dot plots was obtained from 1 fetus. Weights of fetuses are expressed relative to the mean of fetal weights from mock-infected IFNAR^-/- mice.

Kam, Y. W., Lee, C. Y. P., Teo, T. H., Howland, S. W., Amrun, S. N., Lum, F. M., ... & Rénia, L. (2017). Cross-reactive dengue human monoclonal antibody prevents severe pathologies and death from Zika virus infections. JCI insight, 2(8).

Figure 14 Viral load in the (C) amniotic fluid and (D) organs of fetuses from ZIKV-infected IFNAR^-/- pregnant mice receiving isotype, SIgN-3C, and SIgN-3C-LALA antibodies (all groups n ≥ 7).

Each data point in dot plots was obtained from 1 fetus. Mice were inoculated with 107 PFU ZIKV i.v. on E10.5 and treatments were given on days 0, 1, and 3 after infection. Mice were given 0.5 mg of isotype or treatment antibody per dose. All animals were harvested at E16.5. All data were analyzed using the Kruskal-Wallis test with Dunn's multiple comparison. ^*P < 0.05, ^***P < 0.001.

Kam, Y. W., Lee, C. Y. P., Teo, T. H., Howland, S. W., Amrun, S. N., Lum, F. M., ... & Rénia, L. (2017). Cross-reactive dengue human monoclonal antibody prevents severe pathologies and death from Zika virus infections. JCI insight, 2(8).