Department of Pharmacognosy and Pharmaceutical Botany, Faculty of Pharmaceutical Sciences and Research Unit for Plant-Produced Pharmaceuticals, Chulalongkorn University, Bangkok, Thailand.
Faculty of Pharmacy, Mahidol University, Bangkok, Thailand.
Plant Cell Rep. 2021 Apr;40(4):723-733. doi: 10.1007/s00299-021-02670-z. Epub 2021 Feb 13.
Plant expression platform is the new source of immunoglobulin G (IgG) toward small low-molecular-weight targets. The plant-made monoclonal antibody-based immunoassay exhibits comparable analytical performance with hybridoma antibody. Immunoassays for small molecules are efficiently applied for monitoring of serum therapeutic drug concentration, food toxins, environmental contamination, etc. Immunoglobulin G (IgG) is usually produced using hybridoma cells, which requires complicated procedures and expensive equipment. Plants can act as alternative and economic hosts for IgG production. However, the production of free hapten (low-molecular-weight target)-recognizing IgG from plants has not been successfully developed yet. The current study aimed at creating a plant platform as an affordable source of IgG for use in immunoassays and diagnostic tools. The functional IgG was expressed in Nicotiana benthamiana leaves infiltrated with Agrobacterium tumefaciens strain GV3101 with recombinant geminiviral vectors (pBY3R) occupying chimeric anti-miroestrol IgG genes. The appropriate assembly between heavy and light chains was achieved, and the yield of expression was 0.57 µg/g fresh N. benthamiana leaves. The binding characteristics of the IgG to miroestrol and binding specificity to related compounds, such as isomiroestrol and deoxymiroestrol, were similar to those of hybridoma-produced IgG (monoclonal antibody, mAb). The plant-based mAbs exhibited high sensitivity for miroestrol (IC, 23.2 ± 2.1 ng/mL), precision (relative standard deviation ≤ 5.01%), and accuracy (97.8-103% recovery), as determined using quantitative enzyme-linked immunosorbent assay. The validated enzyme-linked immunosorbent assay was applicable to determine miroestrol in plant samples. Overall, the plant-produced functional IgG conserved the binding activity and specificity of the parent IgG derived from mammalian cells. Therefore, the plant expression system may be an efficient and affordable platform for the production of antibodies against low-molecular-weight targets in immunoassays.
植物表达平台是针对小分子低分子量靶标的免疫球蛋白 G (IgG)的新来源。基于植物制造的单克隆抗体的免疫分析与杂交瘤抗体表现出相当的分析性能。用于小分子的免疫分析高效应用于监测血清治疗药物浓度、食物毒素、环境污染等。免疫球蛋白 G (IgG)通常使用杂交瘤细胞生产，这需要复杂的程序和昂贵的设备。植物可以作为 IgG 生产的替代和经济宿主。然而，尚未成功开发从植物中生产游离半抗原（低分子量靶标）识别 IgG。本研究旨在创建一个植物平台，作为用于免疫分析和诊断工具的负担得起的 IgG 来源。在被重组双生病毒载体（pBY3R）占据嵌合抗米雌酚 IgG 基因的农杆菌菌株 GV3101 浸润的 Nicotiana benthamiana 叶片中表达功能性 IgG。实现了重链和轻链之间的适当组装，表达产量为 0.57 µg/g 新鲜的 N. benthamiana 叶片。该 IgG 对米雌酚的结合特性和对相关化合物（如异米雌酚和去氧米雌酚）的结合特异性与杂交瘤产生的 IgG（单克隆抗体，mAb）相似。基于植物的 mAbs 对米雌酚表现出高灵敏度（IC，23.2 ± 2.1 ng/mL）、精密度（相对标准偏差≤5.01%）和准确度（97.8-103%回收率），如使用定量酶联免疫吸附测定法测定。验证的酶联免疫吸附测定法适用于测定植物样品中的米雌酚。总的来说，植物产生的功能性 IgG 保留了源自哺乳动物细胞的亲本 IgG 的结合活性和特异性。因此，植物表达系统可能是用于免疫分析中针对低分子量靶标的抗体生产的高效且经济的平台。
