The 21st century has been particularly productive for the biopharmaceutical industry, with the introduction of several classes of innovative therapeutics, such as monoclonal antibodies and related compounds, gene therapy products, and RNA-based modalities. All these new molecules are susceptible to aggregation and fragmentation, which necessitates a size variant analysis for their comprehensive characterization. Size exclusion chromatography (SEC) is one of the reference techniques that can be applied. The analytical techniques for mAbs are now well established and some of them are now emerging for the newer modalities. In this context, the objective of this review article is: to provide a short historical background on SEC, to suggest some clear guidelines on the selection of packing material and mobile phase for successful method development in modern SEC; and to highlight recent advances in SEC, such as the use of narrow-bore and micro-bore columns, ultra-wide pore columns, and low-adsorption column hardware. Some important innovations, such as recycling SEC, the coupling of SEC with mass spectrometry, and the use of alternative detectors such as charge detection mass spectrometry and mass photometry are also described. In addition, this review discusses the use of SEC in multidimensional setups and shows some of the most recent advances at the preparative scale. In the third part of the article, the possibility of SEC for the characterization of new modalities is also reviewed. The final objective of this review is to provide a clear summary of opportunities and limitations of SEC for the analysis of different biopharmaceutical products.

中文翻译：

---

生物制药产品的尺寸排阻色谱：从当前的蛋白质实践到病毒载体、核酸和脂质纳米粒子的新兴趋势

21 世纪对于生物制药行业来说尤其富有成效，随着几类创新疗法的推出，例如单克隆抗体和相关化合物、基因治疗产品和基于 RNA 的治疗方式。所有这些新分子都容易聚集和断裂，因此需要对其进行尺寸变异分析以进行全面表征。尺寸排阻色谱（SEC）是可以应用的参考技术之一。单克隆抗体的分析技术现已十分成熟，其中一些分析技术正在出现新的模式。在这种背景下，这篇综述文章的目的是：提供 SEC 的简短历史背景，为现代 SEC 成功方法开发的包装材料和流动相的选择提出一些明确的指导方针；并重点介绍 SEC 的最新进展，例如窄孔和微孔色谱柱、超宽孔色谱柱和低吸附色谱柱硬件的使用。还描述了一些重要的创新，例如回收 SEC、SEC 与质谱的耦合以及电荷检测质谱和质量光度测定等替代检测器的使用。此外，本综述讨论了 SEC 在多维设置中的使用，并展示了制备规模的一些最新进展。在文章的第三部分，还回顾了 SEC 表征新模式的可能性。本次审查的最终目标是清晰总结 SEC 分析不同生物制药产品的机会和局限性。