Detection and activation of HIV broadly neutralizing antibody precursor B cells using anti-idiotypes

Bancroft, Tara; DeBuysscher, Blair L.; Weidle, Connor; Schwartz, Allison; Wall, Abigail; Gray, Matthew D.; Feng, Junli; Steach, Holly R.; Fitzpatrick, Kristin S.; Gewe, Mesfin M.; Skog, Patrick D.; Doyle-Cooper, Colleen; Ota, Takayuki; Strong, Roland K.; Nemazee, David; Pancera, Marie; Stamatatos, Leonidas; McGuire, Andrew T.; Taylor, Justin J.

To our knowledge, this study represents the first time antiidiotypes have been used to identify and analyze naive B cells expressing BCRs with similarities to the inferred germline sequence of a broadly neutralizing HIV-1-specific antibody. An important result within these analyses is the finding that different anti-idiotypic antibodies can have different modes of interaction with BCRs similar to the one of interest. The binding profile analysis (Fig. 1 C) and crystal structure data (Fig. 2) suggested that IB2 would select only for BCRs with iglb12-like V H 1-3 + heavy chains, while IB3 would select for BCRs with iglb12-like V H 1-3 + heavy and V K 3-20 + light chains, perhaps as a subset of the IB2-binding population. Sorting and sequencing BCRs from human B cells was therefore important and revealed that IB2 and IB3 selected for completely different populations of cells, with IB2 selecting far more stringently for V H 1-3 + BCRs compared with IB3. Given these findings, we are exploring ways to better predict which anti-idiotypes will be better at identifying BCRs of interest. Nevertheless, our results indicate that Data are pooled from four independent experiments, which were normalized to account for experiment-to-experiment variability by displaying the data as a geometric mean fluorescence intensity (gMFI) fold increase over background fluorescence in CD19 − B220 − non-B cells. The bar indicates the mean, and P values (**, P < 0.001; ***, P < 0.001) were determined using an unpaired two-tailed Student's t test (n = 3-11). The P values (*, P < 0.05; **, P < 0.01; ***, P < 0.001) were determined using an unpaired two-tailed Student's t test.