Figure 3: Identification of CD4-binding site antibodies. | Scientific Reports

Figure 3: Identification of CD4-binding site antibodies.

From: Broad and potent cross clade neutralizing antibodies with multiple specificities in the plasma of HIV-1 subtype C infected individuals

Figure 3

(A) ELISA with RSC3 and RSC3∆371I/P363N recombinant proteins. 12 BCN plasma samples were tested at a dilution of 1:100 to screen for the presence of CD4 binding site directed antibodies. Four of the 12 BCN plasma samples tested demonstrated significantly stronger binding to the wild type RSC3 protein as compared to the mutant protein. (B) RSC3 and RSC3∆371I/P363N specific antibodies were eluted from four BCN plasma samples which revealed the presence of CD4 binding site targeting antibodies (NAB033, NAB059, NAB063 and NAB069) using tosyl activated MyOne Dynabeads. ELISA was again performed with eluted IgG antibody (concentration 10 to 0.0001 μg/ml) with RSC3 and RSC3∆371I/P363N recombinant proteins at 2 μg/ml. Monoclonal antibodies VRC01, VRC03, B12 and 3BNC117 were used as positive controls and HHP as negative control. (C) Neutralization titer (IC50) assay performed with eluted IgG antibody (concentration 10 to 0.0001 μg/ml) against four pseudoviruses and one mutant (JR-FL D279A). VRC01 and b12 MAbs were used as positive control bNabs against CD4BS and MuLV, as negative control.

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