NGS of phage antibody outputs from whole-cell panning. NGS of phage antibody outputs from whole-cell panning

We present a target-unbiased approach for antibody discovery that relies on generating mAbs against native target cell surfaces via phage display. This method combines a previously reported method for improved whole-cell phage display selections with next-generation sequencing analysis to efficiently identify mAbs with the desired target cell reactivity. This approach enabled the identification of three multiple myeloma cell surface antigens, and cognate monoclonal antibody probes. Overall design: Rabbit phage antibody libraries were selected against human multiple myeloma cell lines, HEK293-ROR1-TetOn cells, and healthy donor-derived human PBMCs using the Fab-phage Biotinylation and Capture (FBC) method. Selected antibody outputs were subjected to NGS analysis for HCDR3 profiling.

本研究报道了一种非靶标偏倚性的抗体发现策略，该策略依托噬菌体展示技术，针对天然靶标细胞表面生成单克隆抗体（monoclonal antibody, mAb）。该方法将此前报道的优化全细胞噬菌体展示筛选方案与下一代测序（next-generation sequencing, NGS）分析相结合，可高效筛选出具备预期靶标细胞反应活性的单克隆抗体。本策略成功鉴定出3种多发性骨髓瘤细胞表面抗原及其对应的单克隆抗体探针。

整体实验设计：本研究采用Fab噬菌体生物素化与捕获（Fab-phage Biotinylation and Capture, FBC）方法，针对人多发性骨髓瘤细胞系、HEK293-ROR1-TetOn细胞以及健康供体来源的外周血单个核细胞（peripheral blood mononuclear cell, PBMC）筛选兔噬菌体抗体库。对筛选得到的抗体产物进行下一代测序分析，以完成重链互补决定区3（heavy chain complementarity determining region 3, HCDR3）的谱型分析。