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. 2016 Feb 24;7:10618. doi: 10.1038/ncomms10618

Figure 3. Antibody responses elicited in WT and knock-in gl3BNC60 mice after immunization.

Figure 3

(a) Bone marrow cells (gated on; live cells, CD4−, CD8−, GR-1− and B220+) of naive WT (top) and naive gl3BNC60 knock-in mice (bottom) were stained for IgD and IgM as indicated to identify immature (IgM−/low, IgD−), and mature (IgM+, IgD+) B-cell populations. (b) Splenocytes from naive WT and gl3BNC60 knock-in mice (gated on; live cells, CD4−, CD8−, GR-1−, B220+ and CD93−) were stained with CD21 and CD23 as indicated to identify follicular (CD21low/CD23high) and marginal zone (CD21high/CD23low) B cells. Panels a and b show representative FACS diagrams from one individual mouse of five. (c) Serum IgG collected prior to (naive) and following one (p1) or two immunizations (p2) with 426cTM4ΔV1-V3 gp140 in Alum Imject were tested for binding to 426cTM4ΔV1-V3 gp140 (closed circles) or 426cTM4ΔV1-V3.gp140.D368R.E370A protein (KO; open circles) in WT (left panel, n=3) and knock-in gl3BNC60 mice (right panel, n=3) by ELISA. Lines connecting the black and white circles indicate OD values for 426cTM4ΔV1-V3 gp140 or 426cTM4ΔV1-V3.gp140.D368R.E370A proteins from the same mouse at the indicated concentrations. (d) Same as in c but WT mice (n=5) or knock-in gl3BNC60 (n=5) mice were immunized once with 426cTM4ΔV1-V3 gp140 dextramer in Alum Imject. (e) Same as in c, but WT (n=5) or knock-in gl3BNC60 mice (n=5) were immunized once with 426cTM4ΔV1-V3 gp120-dextramers in Alum Imject (left panel) or in Ribi adjuvant (n=5) (right panel). (f) Same as in c, but knock-in gl3BNC60 mice (n=5) were immunized once with 426c.TM4ΔV1-V3 gp120-C4b in Ribi adjuvant. (g) Same as in c but WT mice (n=3) or knock-in gl3BNC60 mice (n=5) were immunized once with 426cTM4ΔV1-V3 gp120-ferritin in Alum Imject. (h) Serum IgG from WT (n=4) and knock-in 3BNC60 mice (n=5) after one (left panel) or two (right panel) immunizations with WT 426c gp140-dextramer in Alum Imject were tested for binding to WT 426c gp140 (closed squares) or 426c.D368R.E370A (426c-KO, open squares) by ELISA. Lines between black and white squares indicate OD values for WT 426c gp140 or 426c-KO proteins from the same mouse at the indicated concentrations.