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A generalized HIV vaccine design strategy for priming of broadly neutralizing antibody responses.

Science. 2019-12; 
Steichen JM, Lin YC, Havenar-Daughton C, Pecetta S, Ozorowski G, Willis JR, Toy L, Sok D, Liguori A, Kratochvil S, Torres JL, Kalyuzhniy O, Melzi E, Kulp DW, Raemisch S, Hu X, Bernard SM, Georgeson E, Phelps N, Adachi Y, Kubitz M, Landais E, Umotoy J, Robinson A, Briney B, Wilson IA, Burton DR, Ward AB, Crotty S, Batista FD, Schief WR
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Gene Synthesis Mutations found in the most enriched clones were incorporated into the most recent designs, synthesized at Genscript either as C-terminal His tagged gp120s or MD39/MD64 trimers in the pHLsec vector and expressed and purified as described previously (22)...To obtain multivalent immunogens, trimers were genetically fused to ferritin from Helicobacter pylori using a short flexible linker. Genes were codon optimized for HEK293 cells and cloned into the pHLsec plasmid (GenScript)...mAb binding ELISAs were performed by capturing antigen (1 µg/ml) onto plates precoated with anti-His antibody (1 µg/ml; Genscript) and blocked with blocking buffer (5% skim milk, 1% fetal bovine serum, 0.2% tween 20 in PBS)...The capture reagent was His-tag Rabbit pAb (GenScript Cat. No. A00174). Get A Quote
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摘要

Vaccine induction of broadly neutralizing antibodies (bnAbs) to HIV remains a major challenge. Germline-targeting immunogens hold promise for initiating the induction of certain bnAb classes; yet for most bnAbs, a strong dependence on antibody heavy chain complementarity-determining region 3 (HCDR3) is a major barrier. Exploiting ultradeep human antibody sequencing data, we identified a diverse set of potential antibody precursors for a bnAb with dominant HCDR3 contacts. We then developed HIV envelope trimer-based immunogens that primed responses from rare bnAb-precursor B cells in a mouse model and bound a range of potential bnAb-precursor human naïve B cells in ex vivo screens. Our repertoire-guided germline... More

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