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. 2017 Aug 7;214(8):2471–2490. doi: 10.1084/jem.20170633

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© 2017 Sanjuan Nandin et al.

This article is available under a Creative Commons License (Attribution 4.0 International, as described at https://creativecommons.org/licenses/by/4.0/).

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Figure 7. — Generation of gp120-specific antibodies from single sorted plasma cells. (a) The specificity of the gp120 recombinant antibodies generated after stimulating memory B cells from five HIV-uninfected individuals was analyzed by ELISA. Cognate pairs of IgH and IgL genes were introduced into HEK293T cells by transient cotransfection, and culture supernatants were tested for the presence of gp120-specific IgGs. The mean IgG antibody concentration was 1 µg/ml. Cutoff value for a positive signal is taken as more than twofold above background. (b) Pie charts outlining the frequency of the gp120-specific clones. The number in the center of the pie chart indicates the percentage of the gp120-positive antibodies. Numbers outside the pie chart indicate the number of antibody gene sequences with identical IgH and IgL chain rearrangements. (c) Reactivity of the gp120-positive antibodies against a panel of foreign antigens was tested by ELISA. The cutoff value for positivity has been taken as more than twofold background. (d) Octet measurements of the kinetics of selected gp120-reactive antibodies to the recombinant wt 92BR gp-120 are shown. Arrows represent the serial dilution of the antibodies; K_d values are highlighted inside the charts.