抗体药物在疾病诊断中越来越受欢迎,靶向治疗,和免疫预防由于其具有高靶向能力的特点,特异性强,低毒性,和轻微的副作用。对抗体药物的需求稳步增长,生产规模不断扩大。通过单克隆抗体的高容量生产,上游细胞培养技术得到了极大的改善。然而,抗体的下游纯化在生产过程中存在瓶颈。此外,抗体的纯化成本极高,约占抗体生产总成本的50%-80%。色谱技术,鉴于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,和抛光。不同的色谱技术,如亲和色谱法,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,在每个阶段使用。亲和层析,主要是蛋白A亲和层析,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,如离子交换色谱法,疏水相互作用色谱,和混合模式色谱,用于中间体纯化和抗体精制。亲和仿生色谱和疏水电荷诱导色谱可以产生纯度与通过蛋白A色谱获得的抗体相当的抗体,通过采用具有良好选择性的人工化学/短肽配体,高稳定性,和低成本。温度响应色谱是用于分离和纯化抗体的有前途的技术。在这项技术中,通过简单地调节柱温来控制抗体捕获和洗脱,这极大地消除了在酸性洗脱条件下抗体聚集和失活的风险。不同色谱方法的组合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用策略。这篇综述概述了使用色谱法纯化抗体领域的最新进展,并讨论了该技术的未来发展。
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.