School of Public Health, Wannan Medical College, Wuhu, 241002, China.
Hefei Institutes of Physical Science, Chinese Academy of Sciences, Hefei, 230031, China.
BMC Biotechnol. 2023 Jul 28;23(1):24. doi: 10.1186/s12896-023-00798-2.
Biological laboratories and companies involved in antibody development need convenient and versatile methods to detect highly active antibodies.
To develop a mammalian cell-based ZZ display system for antibody quantification, the eukaryotic ZZ-displayed plasmid was constructed and transfected into CHO cells. After screening by flow cytometric sorting, the stable ZZ display cells were incubated with reference IgG and samples with unknown IgG content for 40 min at 4℃, the relative fluorescence intensity of cells was analyzed and the concentration of IgG was calculated.
By investigating the effects of different display-associated genetic elements, a eukaryotic ZZ-displaying plasmid with the highest display efficiency were constructed. After transfection and screening, almost 100% of the cells were able to display the ZZ peptide (designated CHO-ZZ cells). These stable CHO-ZZ cells were able to capture a variety of IgG, including human, rabbit, donkey and even mouse and goat. CHO-ZZ cells could be used to quantify human IgG in the range of approximately 12.5-1000 ng/mL, and to identify high-yielding engineered monoclonal cell lines.
We have established a highly efficient CHO-ZZ display system in this study, which enables the quantification of IgG from various species under physiological conditions. This system offers the advantage of eliminating the need for antibody purification and will contribute to antibody development.
从事抗体开发的生物实验室和公司需要方便、通用的方法来检测高活性抗体。
为了开发一种用于抗体定量的基于哺乳动物细胞的 ZZ 展示系统，构建了真核 ZZ 展示质粒并转染 CHO 细胞。通过流式细胞分选筛选后，将稳定的 ZZ 展示细胞与参比 IgG 和未知 IgG 含量的样品在 4℃下孵育 40min，分析细胞的相对荧光强度，并计算 IgG 的浓度。
通过研究不同展示相关遗传元件的影响，构建了展示效率最高的真核 ZZ 展示质粒。转染和筛选后，几乎 100%的细胞能够展示 ZZ 肽（命名为 CHO-ZZ 细胞）。这些稳定的 CHO-ZZ 细胞能够捕获各种 IgG，包括人、兔、驴，甚至鼠和山羊的 IgG。CHO-ZZ 细胞可用于定量约 12.5-1000ng/mL 范围内的人 IgG，并鉴定高产工程单克隆细胞系。
本研究建立了一种高效的 CHO-ZZ 展示系统，可在生理条件下定量检测来自不同物种的 IgG。该系统具有无需抗体纯化的优势，将有助于抗体开发。
