抗体经木瓜蛋白酶消化后，V(H)3 Fab与Fc的蛋白A分离得到改善。
Improved Protein-A separation of V(H)3 Fab from Fc after papain digestion of antibodies.
作者信息
Seldon Therese A, Hughes Karen E, Munster David J, Chin David Y, Jones Martina L
机构信息
Mater Medical Research Institute, South Brisbane, Australia.
出版信息
J Biomol Tech. 2011 Jul;22(2):50-2.
Abstract
Antibody-binding fragments (Fab) are generated from whole antibodies by treatment with papain and can be separated from the Fc component using Protein-A affinity chromatography. Commercial kits are available, which facilitate the production and purification of Fab fragments; however, the manufacturer fails to report that this method is inefficient for antibodies with V(H)3 domains as a result of the intrinsic variable region affinity for Protein-A. A commercially available, modified Protein-A resin (MabSelect SuRe) has been engineered for greater stability. Here, we report that an additional consequence of the modified resin is the ability to purify V(H)3 family Fab fragments, which cannot be separated effectively from other components of the papain digest by traditional Protein-A resin. This improvement of a commonly used procedure is of significance, as increasingly, therapeutic antibodies are being derived from human origin, where V(H)3 is the most abundantly used variable region family.
摘要
抗体结合片段（Fab）是通过用木瓜蛋白酶处理完整抗体产生的，并且可以使用蛋白A亲和色谱法与Fc成分分离。有商业试剂盒可供使用，这便于Fab片段的生产和纯化；然而，制造商未报告由于可变区对蛋白A的固有亲和力，该方法对具有V(H)3结构域的抗体效率低下。一种市售的改良蛋白A树脂（MabSelect SuRe）已被设计用于提高稳定性。在此，我们报告改良树脂的另一个结果是能够纯化V(H)3家族Fab片段，而传统蛋白A树脂无法将其与木瓜蛋白酶消化产物的其他成分有效分离。这种常用方法的改进具有重要意义，因为越来越多的治疗性抗体源自人类，其中V(H)3是使用最广泛的可变区家族。
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