纳米抗体筛选技术研究进展

Advances in nanobody screening technology

摘要:

纳米抗体(nanobody, Nb)是在骆驼科血清中发现的一种新型抗体，具有体积小、特异性强、稳定性高、易于表达和能识别隐藏的抗原表位等优势，在各个领域具有广泛的应用价值。本文介绍了纳米抗体筛选与优化过程，包括纳米抗体文库构建、体外展示和亲和力成熟3个重要技术阶段的分类与特点。其中，简要描述了天然、免疫及半合成/合成文库的制备方法与重要参数，并系统介绍了应用噬菌体、酵母、细菌、核糖体/mRNA和真核细胞等表面展示系统，以及酵母双杂交、高通量测序和质谱鉴定方法，共8种不同体外展示技术进行快速筛选的方法及其优缺点，汇总用于提升纳米抗体功能可靠性的体外及计算机辅助亲和力成熟技术平台，为综合运用各种技术手段快速获得稳定、可靠、特异的纳米抗体类药物或诊断制剂提供了参考。

Abstract:

Nanobody (Nb) is a novel type of antibody discovered in the serum of Camelidae. It is characterized by its small size, high specificity, stability, and ease of preparation. Nanobodies exhibit the ability to identify hidden epitopes and have diverse applications across various fields. This review aims to introduce three key stages in the screening and optimization of nanobodies, including nanobody library construction, in vitro surface display, and affinity maturation. We provide a brief description of preparation and characteristics of natural, immunological, and semi-synthetic/synthetic libraries. Additionally, we systematically explain eight in vitro display methods, including phage display, yeast display, bacterial display, ribosome display/mRNA display, and eukaryotic cell display. Furthermore, we discuss the application of yeast two-hybrid system high-throughput sequencing and mass spectrometry identification. A thorough analysis of their advantages and limitations is presented in this protocols. Finally, we summarize the platforms for in vitro or computer-aided affinity maturation techniques aimed at enhancing the functional stability of nanobodies. Consequently, this review provides a comprehensive approach to the integrated utilization of various technologies for the rapid development of stable, reliable, and specific nanobody-based drugs or diagnostic agents.