Highly efficient antibody purification with controlled orientation of protein A on magnetic nanoparticles,RSC Medicinal Chemistry

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Highly efficient antibody purification with controlled orientation of protein A on magnetic nanoparticles
RSC Medicinal Chemistry ( IF 4.1 ) Pub Date : 2017-11-02 00:00:00 , DOI: 10.1039/c7md00468k
Sunghyun Kim _{1,

2,

3} , Daekyung Sung _{1,

2,

3} , Jeong Ho Chang _{1,

2,

3}

Affiliation

In this study, we prepared protein A grafted magnetic nanoparticles for the industrial large-scale purification of antibodies with enhancement of binding capacity and immobilization by controlled orientation with chlorophenylsilane (CPTMS) on the surface. For site-specific immobilization of protein A, genetically modified protein A with a cysteine residue was expressed in E. coli and purified by affinity chromatography. To improve the surface area to volume ratio and increase the immobilization amount of protein A, chlorophenylsilane functionalized magnetic nanoparticles (CPTMS@MNPs) were prepared, which are smaller nanoparticles with an average diameter of 20 nm compared to commercial magnetic microparticles (Dynabeads) with an average size of 2.8 μm. The CPTMS@MNPs showed the enhancement of protein A immobilization and binding capacity to antibodies, being 11.5-fold and 7-fold higher than those of commercial Dynabeads, respectively. In addition, the CPTMS@MNPs retained about 80% of the initial protein binding capacity until the third stage of recycling. Therefore, protein A grafted CPTMS@MNPs may be useful for the industrial large-scale purification of antibodies.

中文翻译：

高效的抗体纯化，可控制蛋白A在磁性纳米颗粒上的定向

在这项研究中，我们制备了蛋白A接枝的磁性纳米颗粒，用于工业化大规模纯化抗体，具有增强的结合能力，并通过在表面上与氯苯基硅烷（CPTMS）受控地定向来固定化。对于蛋白质A的位点特异性固定，在大肠杆菌中表达了带有半胱氨酸残基的基因修饰的蛋白质A。并通过亲和色谱法纯化。为了提高表面积与体积之比并增加蛋白A的固定化量，制备了氯苯基硅烷官能化的磁性纳米粒子（CPTMS @ MNPs），与商用磁性微粒（Dynabeads）相比，它们是平均直径为20 nm的较小纳米粒子。平均尺寸为2.8μm。CPTMS @ MNPs显示出增强的蛋白A固定和对抗体的结合能力，分别比市售Dynabeads高11.5倍和7倍。此外，CPTMS @ MNP保留了约80％的初始蛋白质结合能力，直到回收的第三阶段。因此，蛋白A嫁接的CPTMS @ MNPs可用于抗体的工业大规模纯化。

更新日期：2017-11-27

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