Development of a novel capture step for purification of antigen binding fragments (Fabs)

IF 1.4 4区生物学 Q4 BIOCHEMICAL RESEARCH METHODS

Protein expression and purification Pub Date : 2024-12-17 DOI:10.1016/j.pep.2024.106647

Anupa Anupa , Anurag S. Rathore

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引用次数: 0

Abstract

Antigen binding fragments (Fabs) are an emerging class of biotherapeutics, widely accepted as an alternative to the traditional monoclonal antibodies (mAbs). The small size of the Fabs offers better tissue penetrability and lack of Fc region, thereby resulting in reduced side effects. However, since Fab molecules lack Fc region, Protein A chromatography (the ubiquitous capture step in mAb platforms) cannot be employed. Conventional Fab purification platforms employ affinity-based Protein L chromatography for capture, which is effective but just like Protein A an expensive step to use in commercial production. In this paper, we propose use of salt precipitation followed by hydrophobic interaction chromatography (HIC) as a capture chromatography step for Fabs. The precipitation step enhanced the purity from 30 % to 65 % and the HIC chromatography step further raising it to 80 % (by RP-HPLC) with a combined process recovery of greater than 95 %. The proposed capture process offers a substantial advantage with respect to the cost of goods when compared to the traditional, affinity-based approaches.

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抗原结合片段（fab）纯化新捕获步骤的开发。

抗原结合片段（fab）是一类新兴的生物治疗药物，被广泛接受为传统单克隆抗体（mab）的替代品。晶圆片的小尺寸提供了更好的组织穿透性和缺乏Fc区，从而减少了副作用。然而，由于Fab分子缺乏Fc区，因此无法采用蛋白A层析（mAb平台中普遍存在的捕获步骤）。传统的Fab纯化平台采用基于亲和的蛋白质L层析来捕获，这是有效的，但就像蛋白质A一样，在商业生产中使用是一个昂贵的步骤。在本文中，我们建议使用盐沉淀然后疏水相互作用色谱（HIC）作为fab的捕获色谱步骤。沉淀步骤将纯度从30%提高到65%，高效液相色谱步骤将纯度进一步提高到80% (RP-HPLC)，联合工艺回收率大于95%。与传统的基于亲和力的方法相比，拟议的捕获过程在货物成本方面具有实质性优势。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Protein expression and purification 生物-生化研究方法

CiteScore

3.70

自引率

6.20%

发文量

120

审稿时长

32 days

期刊介绍： Protein Expression and Purification is an international journal providing a forum for the dissemination of new information on protein expression, extraction, purification, characterization, and/or applications using conventional biochemical and/or modern molecular biological approaches and methods, which are of broad interest to the field. The journal does not typically publish repetitive examples of protein expression and purification involving standard, well-established, methods. However, exceptions might include studies on important and/or difficult to express and/or purify proteins and/or studies that include extensive protein characterization, which provide new, previously unpublished information.