Department of Pharmacology, Kobe Pharmaceutical University.
Research Institute for Production Development.
Biol Pharm Bull. 2022;45(8):1022-1026. doi: 10.1248/bpb.b22-00220.
The emu is the second largest ratite; thus, their sera and egg yolks, obtained after immunization, could provide therapeutic and diagnostically important immunoglobulins with improved production efficiency. Reliable purification tools are required to establish a pipeline for supplying practical emu-derived antibodies, the majority of which belongs to the immunoglobulin Y (IgY) class. Therefore, we generated a monoclonal secondary antibody specific to emu IgY. Initially, we immunized an emu with bovine serum albumin multiply haptenized with 2,4-dinitrophenyl (DNP) groups. Polyclonal emu anti-DNP antibodies were partially purified using conventional precipitation method and used as antigen for immunizing a BALB/c mouse. Splenocytes were fused with myeloma cells and a hybridoma clone secreting a desirable secondary antibody (mAb#2-16) was established. The secondary antibody bound specifically to emu-derived IgY, distinguishing IgYs from chicken, duck, ostrich, quail, and turkey, as well as human IgGs. Affinity columns immobilizing the mAb#2-16 antibodies enabled purification of emu IgY fractions from sera and egg yolks via simple protocols, with which we succeeded in producing IgYs specific to the severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) spike protein with a practical binding ability. We expect that the presented purification method, and the secondary antibody produced in this study, will facilitate the utilization of emus as a novel source of therapeutic and diagnostic antibodies.
鸸鹋是第二大平胸鸟类；因此，它们的血清和蛋黄在免疫后可以提供具有改进生产效率的治疗和诊断上重要的免疫球蛋白。需要可靠的纯化工具来建立供应实用鸸鹋衍生抗体的管道，其中大多数属于免疫球蛋白 Y（IgY）类。因此，我们生成了一种针对鸸鹋 IgY 的单克隆二次抗体。最初，我们用 2,4-二硝基苯（DNP）基团多缀合的牛血清白蛋白免疫鸸鹋。使用常规沉淀法部分纯化多克隆鸸鹋抗-DNP 抗体，并将其用作免疫 BALB/c 小鼠的抗原。脾细胞与骨髓瘤细胞融合，并建立了分泌理想二次抗体（mAb#2-16）的杂交瘤克隆。该二次抗体特异性结合鸸鹋源性 IgY，可区分来自鸡、鸭、鸵鸟、鹌鹑和火鸡以及人 IgG 的 IgY。用 mAb#2-16 抗体固定化的亲和柱可通过简单的方案从血清和蛋黄中纯化鸸鹋 IgY 级分，我们成功地用其生产了具有实用结合能力的针对严重急性呼吸综合征冠状病毒 2（SARS-CoV-2）刺突蛋白的特异性 IgY。我们期望所提出的纯化方法和本研究中产生的二次抗体将促进将鸸鹋用作治疗和诊断抗体的新型来源。
