Bioproduction Research Institute, National Institute of Advanced Industrial Science and Technology (AIST), Central 6, 1-1-1 Higashi, Tsukuba, Ibaraki, 305-8566, Japan.
NGK Spark Plug-AIST Healthcare ・ Materials Cooperative Research Laboratory, National Institute of Advanced Industrial Science and Technology (AIST), 2266-98 Anagahora, Shimoshidami, Moriyama-ku, Nagoya, 463-8560, Japan.
Sci Rep. 2021 Feb 9;11(1):3233. doi: 10.1038/s41598-021-82457-0.
Here, we describe porous zirconia particles (PZPs) optimized for the purification of immunoglobulins. PZPs, with a pore size of approximately 10 nm, were designed to specifically interact with antibodies via surface modification with a phosphate functional group. A simple PZP purification method based on precipitation enabled efficient purification of mouse anti-glycosphingolipid globoside/Gb4Cer monoclonal IgM (κ-light chains) from hybridoma culture supernatants. Over 99% of contaminating proteins were removed by the PZP purification process, and approximately 50% of the IgM was recovered in the purified fraction after eluting the PZP-adsorbed antibodies with 100 mM phosphate buffer. Other IgG3 and IgM monoclonal antibodies that react with Gb4Cer or α2,6-sialyl LacNAc-modified glycoproteins could also be purified using PZPs and elution buffer at concentrations of 100-500 mM. All of the purified antibodies retained their antigen reactivity and specificity, indicating that PZP purification does not affect antibody function. As PZP purification is also suitable for purification of IgM consisting of λ-light chains and IgG derived from other mammalian species, it is expected to be applied to the purification of a variety of antibodies, including anti-glycoconjugate IgMs.
在这里，我们描述了优化用于免疫球蛋白纯化的多孔氧化锆颗粒（PZPs）。PZPs 的孔径约为 10nm，通过用磷酸官能团进行表面修饰，旨在与抗体特异性相互作用。基于沉淀的简单 PZP 纯化方法能够从杂交瘤培养上清液中有效地纯化鼠抗神经节苷脂Globoside/Gb4Cer 单克隆 IgM（κ 轻链）。通过 PZP 纯化过程去除了超过 99%的污染蛋白，并且在用 100mM 磷酸盐缓冲液洗脱 PZP 吸附的抗体后，大约 50%的 IgM 被回收在纯化部分中。使用 PZPs 和洗脱缓冲液，其他与 Gb4Cer 或α2,6-唾液酸化 LacNAc 修饰的糖蛋白反应的 IgG3 和 IgM 单克隆抗体也可以在 100-500mM 的浓度下进行纯化。所有纯化的抗体均保留其抗原反应性和特异性，表明 PZP 纯化不影响抗体功能。由于 PZP 纯化也适用于由 λ 轻链组成的 IgM 和源自其他哺乳动物物种的 IgG 的纯化，预计它将应用于各种抗体的纯化，包括抗糖缀合物 IgM。
