Sohyaku. Innovative Research Department, Mitsubishi Tanabe Pharma Corporation, Yokohama, Japan.
Development Department, Tanabe Research Laboratories U.S.A. Inc., San Diego, USA.
Sci Rep. 2021 Jan 19;11(1):1827. doi: 10.1038/s41598-021-81366-6.
Protein A affinity chromatography has been widely used for both laboratory scale purification and commercial manufacturing of monoclonal antibodies and Fc-fusion proteins. Protein A purification is specific and efficient. However, there still remain several issues to be addressed, such as incomplete clearance of impurities including host cell proteins, DNA, aggregates, etc. In addition, the effects of wash buffers in protein A purification on the physicochemical characteristics of antibodies have yet to be fully understood. Here we found a new purification protocol for monoclonal antibodies that can improve physicochemical properties of monoclonal antibodies simply by inserting an additional wash step with a basic buffer after the capture step to the conventional protein A purification. The effects of the alkaline wash on monoclonal antibodies were investigated in terms of physicochemical characteristics, yields, and impurity clearance. The simple insertion of an alkaline wash step resulted in protection of antibodies from irreversible aggregation, reduction in free thiols and impurities, an improvement in colloidal and storage stability, and enhanced yields. This new procedure is widely applicable to protein A affinity chromatography of monoclonal antibodies.
Protein A 亲和层析已广泛应用于单克隆抗体和 Fc 融合蛋白的实验室规模纯化和商业生产。Protein A 纯化具有特异性和高效性。然而，仍有几个问题需要解决，例如宿主细胞蛋白、DNA、聚集物等杂质不完全清除。此外，Protein A 纯化中洗涤缓冲液对抗体理化性质的影响尚未得到充分理解。在这里，我们发现了一种新的单克隆抗体纯化方案，通过在常规 Protein A 纯化的捕获步骤后插入一个碱性缓冲液的额外洗涤步骤，可以简单地改善单克隆抗体的理化性质。从理化特性、产量和杂质清除等方面研究了碱性洗涤对单克隆抗体的影响。简单地插入碱性洗涤步骤可防止抗体发生不可逆聚集，减少游离巯基和杂质，提高胶体和储存稳定性，并提高产量。这种新方法广泛适用于单克隆抗体的 Protein A 亲和层析。
