Human Anti-EBOV GP Recombinant Antibody (clone ma-B06) (CAT#: FAMAB-0526WJ)

Figure 1 Neutralization of vaccination-induced macaque monoclonal antibodies (mAbs).

Bar chart representing the values of half-maximal inhibitory concentration ([IC50] μg/mL) calculated after in vitro neutralization assay using Zaire ebolavirus (EBOV) glycoprotein (GP)-pseudotyped-virus performed with the 14 mAbs that were cloned. In gray are negative controls VRC01 and mAb114 used as a reference. Seven of 14 mAbs showed ability to neutralize in vitro.

Cagigi, A., Misasi, J., Ploquin, A., Stanley, D. A., Ambrozak, D., Tsybovsky, Y., ... & Sullivan, N. J. (2018). Vaccine generation of protective Ebola antibodies and identification of conserved B-cell signatures. The Journal of Infectious Diseases, 218(suppl_5), S528-S536.

Figure 2 Neutralization of vaccination-induced macaque monoclonal antibodies (mAbs).

Dot plots show the-log half-maximal effective concentration ([EC50] μg/mL) representing binding of the 7 neutralizers identified in A to GP (filled circles) and GP_ΔMUC (empty circles). All of the mAbs tested bound GP_ΔMUC with greater potency compared with GP (P = .0097).

Cagigi, A., Misasi, J., Ploquin, A., Stanley, D. A., Ambrozak, D., Tsybovsky, Y., ... & Sullivan, N. J. (2018). Vaccine generation of protective Ebola antibodies and identification of conserved B-cell signatures. The Journal of Infectious Diseases, 218(suppl_5), S528-S536.

Figure 3 Immunoprecipitation of the 7 neutralizers performed with GP_THL.

Four of seven mAbs were able to precipitate GP_THL. As control, input GP_THL is shown on a Coomassie-stained gel.

Cagigi, A., Misasi, J., Ploquin, A., Stanley, D. A., Ambrozak, D., Tsybovsky, Y., ... & Sullivan, N. J. (2018). Vaccine generation of protective Ebola antibodies and identification of conserved B-cell signatures. The Journal of Infectious Diseases, 218(suppl_5), S528-S536.

Figure 4 Western blot on the 4 mAbs identified in C performed with GP_ΔMUC and GP_THL.

For mAb114, none of the mAbs tested were able to bind GP_THL by Western blot. As a positive control for GP_THL, binding of polyclonal serum is shown.

Cagigi, A., Misasi, J., Ploquin, A., Stanley, D. A., Ambrozak, D., Tsybovsky, Y., ... & Sullivan, N. J. (2018). Vaccine generation of protective Ebola antibodies and identification of conserved B-cell signatures. The Journal of Infectious Diseases, 218(suppl_5), S528-S536.

Figure 5 Bar chart representing the percentage of binding inhibition of biotinylated mAb114 to GP_ΔMUC in the presence of each of the 4 putative mAb114-like antibodies.

All mAbs showed a high degree of competition with mAb114 for binding to GP_ΔMUC (calculated at the concentration of 8 μg/mL). The gray bar shows the maximal degree of self-competition of mAb114.

Cagigi, A., Misasi, J., Ploquin, A., Stanley, D. A., Ambrozak, D., Tsybovsky, Y., ... & Sullivan, N. J. (2018). Vaccine generation of protective Ebola antibodies and identification of conserved B-cell signatures. The Journal of Infectious Diseases, 218(suppl_5), S528-S536.

Figure 6 Bar chart on the binding kinetics of the GP1 core antibodies compared with mAb114.

log (KD) values were calculated by biolayer interferometry performed with Fab binding to GP_ΔMUC at neutral and low pH (black and dark gray bars, respectively) as well as to GP_THL at low pH only (light gray bars). All monoclonal antibodies (mAbs) showed similar kinetics compared with mAb114 with ma-D08 showing slightly lower values. Fab fragments derived from papain digestion of mAbs were used for measuring kinetics. This experiment was performed twice. Values are shown from 1 representative experiment.

Cagigi, A., Misasi, J., Ploquin, A., Stanley, D. A., Ambrozak, D., Tsybovsky, Y., ... & Sullivan, N. J. (2018). Vaccine generation of protective Ebola antibodies and identification of conserved B-cell signatures. The Journal of Infectious Diseases, 218(suppl_5), S528-S536.

Figure 7 Bar chart on competition of the GP1 core antibodies with NPC1-dC compared with mAb114 performed by biolayer interferometry using whole IgG binding to GP_THL.

All mAbs showed similar competition with NPC1-dC for binding to GP_THL. Human immunodeficiency virus GP120 (indicated as "Control") and 2 additional mAbs (KZ52 and mAb100) that are known to bind the base of GP_THL far from the receptor binding domain (RBD) were used as negative controls. NPC1-dC was used to assess the maximal degree of self-competition. This experiment was also performed twice, and values are shown from 1 representative experiment.

Cagigi, A., Misasi, J., Ploquin, A., Stanley, D. A., Ambrozak, D., Tsybovsky, Y., ... & Sullivan, N. J. (2018). Vaccine generation of protective Ebola antibodies and identification of conserved B-cell signatures. The Journal of Infectious Diseases, 218(suppl_5), S528-S536.

Figure 8 Electron microscopy reveals that all of the RBD blockers approach glycoprotein (GP) vertically but with a slightly more open angle compared with mAb114 (Fab indicated with green stars).

The scale bars are 10 nm.

Cagigi, A., Misasi, J., Ploquin, A., Stanley, D. A., Ambrozak, D., Tsybovsky, Y., ... & Sullivan, N. J. (2018). Vaccine generation of protective Ebola antibodies and identification of conserved B-cell signatures. The Journal of Infectious Diseases, 218(suppl_5), S528-S536.