Liu Jia-Wei, Tang Chang-Wei, Xia Yi-Ran, Bai Quan
Key Laboratory of Synthetic and Natural Functional Molecule of the Ministry of Education, Institute of Modern Separation Science, Key Laboratory of Modern Separation Science in Shaanxi Province, College of Chemistry & Materials Science, Northwest University, Xi'an 710127, China.
Se Pu. 2024 Jun;42(6):533-543. doi: 10.3724/SP.J.1123.2023.12010.
Antibody drugs are becoming increasingly popular in disease diagnosis, targeted therapy, and immunoprevention owing to their characteristics of high targeting ability, strong specificity, low toxicity, and mild side effects. The demand for antibody drugs is steadily increasing, and their production scale is expanding. Upstream cell culture technology has been greatly improved by the high-capacity production of monoclonal antibodies. However, the downstream purification of antibodies presents a bottleneck in the production process. Moreover, the purification cost of antibodies is extremely high, accounting for approximately 50%-80% of the total cost of antibody production. Chromatographic technology, given its selectivity and high separation efficiency, is the main method for antibody purification. This process usually involves three stages: antibody capture, intermediate purification, and polishing. Different chromatographic techniques, such as affinity chromatography, ion-exchange chromatography, hydrophobic interaction chromatography, mixed-mode chromatography, and temperature-responsive chromatography, are used in each stage. Affinity chromatography, mainly protein A affinity chromatography, is applied for the selective capture and purification of antibodies from raw biofluids or harvested cell culture supernatants. Other chromatographic techniques, such as ion-exchange chromatography, hydrophobic interaction chromatography, and mixed-mode chromatography, are used for intermediate purification and antibody polishing. Affinity biomimetic chromatography and hydrophobic charge-induction chromatography can produce antibodies with purities comparable with those obtained through protein A chromatography, by employing artificial chemical/short peptide ligands with good selectivity, high stability, and low cost. Temperature-responsive chromatography is a promising technique for the separation and purification of antibodies. In this technique, antibody capture and elution is controlled by simply adjusting the column temperature, which greatly eliminates the risk of antibody aggregation and inactivation under acidic elution conditions. The combination of different chromatographic methods to improve separation selectivity and achieve effective elution under mild conditions is another useful strategy to enhance the yield and quality of antibodies. This review provides an overview of recent advances in the field of antibody purification using chromatography and discusses future developments in this technology.
抗体药物因其具有高靶向性、强特异性、低毒性和轻微副作用等特点，在疾病诊断、靶向治疗和免疫预防中越来越受欢迎。对抗体药物的需求稳步增长，其生产规模也在不断扩大。上游细胞培养技术因单克隆抗体的高产量生产而得到了极大改进。然而，抗体的下游纯化是生产过程中的一个瓶颈。此外，抗体的纯化成本极高，约占抗体生产成本的50%-80%。色谱技术因其选择性和高分离效率，是抗体纯化的主要方法。这个过程通常包括三个阶段：抗体捕获、中间纯化和精制。每个阶段使用不同的色谱技术，如亲和色谱、离子交换色谱、疏水相互作用色谱、混合模式色谱和温度响应色谱。亲和色谱，主要是蛋白A亲和色谱，用于从原始生物流体或收获的细胞培养上清液中选择性捕获和纯化抗体。其他色谱技术，如离子交换色谱、疏水相互作用色谱和混合模式色谱，用于中间纯化和抗体精制。亲和仿生色谱和疏水电荷诱导色谱可以通过使用具有良好选择性、高稳定性和低成本的人工化学/短肽配体，生产出纯度与通过蛋白A色谱获得的抗体相当的抗体。温度响应色谱是一种很有前途的抗体分离纯化技术。在这项技术中，只需调节柱温就能控制抗体的捕获和洗脱，这大大消除了在酸性洗脱条件下抗体聚集和失活的风险。结合不同的色谱方法以提高分离选择性并在温和条件下实现有效洗脱，是提高抗体产量和质量的另一种有效策略。本文综述了色谱法在抗体纯化领域的最新进展，并讨论了该技术的未来发展。
