A high-throughput capillary isoelectric focusing immunoassay for fingerprinting protein sialylation

Markely Lam Raga Anggara; Cheung Lila; Choi Young Jun; Estes Scott; Prajapati Shashi; Turyan Iva; Frenkel Ruth; Sosic Zoran; Lambropoulos James; Tescione Lia; Ryll Thomas; Berman Melissa

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A high-throughput capillary isoelectric focusing immunoassay for fingerprinting protein sialylation

机译：用于指纹蛋白唾液酸的高通量毛细管等电聚焦免疫测定

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The serum half-life, biological activity, and solubility of many recombinant glycoproteins depend on their sialylation. Monitoring glycoprotein sialylation during cell culture manufacturing is, therefore, critical to ensure product efficacy and safety. Here a high-throughput method for semi-quantitative fingerprinting of glycoprotein sialylation using capillary isoelectric focusing immunoassay on NanoPro (Protein Simple) platform was developed. The method was specific, sensitive, precise, and robust. It could analyze 2 L of crude cell culture samples without protein purification, and could automatically analyze from 8 samples in 4 h to 96 samples in 14 h without analyst supervision. Furthermore, its capability to detect various changes in sialylation fingerprints during cell culture manufacturing process was indispensable to ensure process robustness and consistency. Moreover, the changes in the sialylation fingerprints analyzed by this method showed strong correlations with intact mass analysis using liquid chromatography and mass spectrometry. (c) 2015 American Institute of Chemical Engineers Biotechnol. Prog., 32:235-241, 2016

机译：许多重组糖蛋白的血清半衰期，生物活性和溶解度取决于它们的唾液酸化。因此，在细胞培养制造过程中监测糖蛋白唾液酸化是至关重要的，以确保产品疗效和安全性至关重要。这里开发了一种使用毛细管等电聚焦免疫测定在纳米孔（蛋白质简单）平台上使用毛细管异电蛋白唾液酸蛋白唾液酸糖的高通量方法。该方法是特异性的，敏感的，精确的和坚固的。它可以分析2L的粗细胞培养样品，没有蛋白质纯化，并且可以在14小时内自动分析在4小时至96个样品中，没有分析人员监督。此外，其能够检测细胞培养制造过程中唾液酸化指纹的各种变化是必不可少的，以确保过程鲁棒性和一致性。此外，该方法分析的唾液酸化指纹的变化显示出使用液相色谱和质谱法的完整质量分析的强相关性。（c）2015美国化学工程研究所生物科技。 Prog。，32：235-241,2016

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来源
《Biotechnology Progress》 |2016年第1期|共7页
作者
Markely Lam Raga Anggara; Cheung Lila; Choi Young Jun; Estes Scott; Prajapati Shashi; Turyan Iva; Frenkel Ruth; Sosic Zoran; Lambropoulos James; Tescione Lia; Ryll Thomas; Berman Melissa;
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作者单位

Biogen Inc Cell Culture Dev High Throughput Analyt Grp 125 Broadway Cambridge MA 02142 USA;

Biogen Inc Cell Culture Dev High Throughput Analyt Grp 125 Broadway Cambridge MA 02142 USA;

Biogen Inc Cell Culture Dev High Throughput Analyt Grp 125 Broadway Cambridge MA 02142 USA;

Biogen Inc Cell Culture Dev High Throughput Analyt Grp 125 Broadway Cambridge MA 02142 USA;

Biogen Inc Cell Culture Dev High Throughput Analyt Grp 125 Broadway Cambridge MA 02142 USA;

Biogen Inc Analyt Dev High Throughput Analyt Grp 125 Broadway Cambridge MA 02142 USA;

Biogen Inc Analyt Dev High Throughput Analyt Grp 125 Broadway Cambridge MA 02142 USA;

Biogen Inc Analyt Dev High Throughput Analyt Grp 125 Broadway Cambridge MA 02142 USA;

Biogen Inc Cell Culture Dev 125 Broadway Cambridge MA 02142 USA;

Biogen Inc Cell Culture Dev 125 Broadway Cambridge MA 02142 USA;

Biogen Inc Cell Culture Dev 125 Broadway Cambridge MA 02142 USA;

Biogen Inc Biomol &

Small Mol Sci 125 Broadway Cambridge MA 02142 USA;

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原文格式 PDF
正文语种 eng
中图分类生物科学;
关键词
carbohydrate; sialic acid; glycoprotein; cell culture; process monitoring;

机译：碳水化合物;唾液酸;糖蛋白;细胞培养;过程监测;

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专利

1. A high-throughput capillary isoelectric focusing immunoassay for fingerprinting protein sialylation [J] . Markely Lam Raga Anggara, Cheung Lila, Choi Young Jun, Biotechnology Progress . 2016,第1期

机译：用于指纹蛋白唾液酸的高通量毛细管等电聚焦免疫测定
2. Capillary isoelectric-focusing immunoassays to study dynamic oncoprotein phosphorylation and drug response to targeted therapies in non-small cell lung cancer [J] . ChenJ.-Q., LeeJ.-H., HerrmannM.A., Molecular cancer therapeutics . 2013,第11期

机译：毛细管等电聚焦免疫测定研究非小细胞肺癌中动态癌蛋白磷酸化和靶向治疗的药物反应
3. Estimation of isoelectric points of human plasma proteins employing capillary isoelectric focusing and peptide isoelectric point markers [J] . Jin Y., Luo GA., Oka T., Electrophoresis: The Official Journal of the International Electrophoresis Society . 2002,第19期

机译：利用毛细管等电聚焦和肽等电点标记估算人血浆蛋白的等电点
4. Online Membrane-Assisted Buffer exchanger coupled with multijunction capillary Isoelectric focusing device enables fractionation of intact human plasma proteins by pl [C] . Mohammad Pirmoradian, Juan Astorga-Wells, Roman A. Zubarev ASMS Conference on Mass Spectrometry and Allied Topics . 2015

机译：与多结毛细管等电聚焦装置联接的在线膜辅助缓冲交换器能够通过PL分馏完整的人血浆蛋白
5. Investigation of protein-drug interaction using capillary isoelectric focusing with whole column imaging detection and spectroscopic techniques . [D] . Mukria, Tibebe Lemma. 2009

机译：使用毛细管等电聚焦结合全柱成像检测和光谱技术研究蛋白质-药物相互作用。
6. Capillary isoelectric-focusing immunoassays to study dynamic oncoprotein phosphorylation and drug response to targeted therapies in non-small cell lung cancer [O] . Jin-Qiu Chen, Jih-Hsiang Lee, Michelle A. Herrmann, -1

机译：毛细管等电聚焦免疫测定研究非小细胞肺癌中动态癌蛋白磷酸化和靶向治疗的药物反应
7. Capillary Isoelectric-Focusing Immunoassays to Study Dynamic Oncoprotein Phosphorylation and Drug Response to Targeted Therapies in Non–Small Cell Lung Cancer [O] . Jin-Qiu Chen, Jih-Hsiang Lee, Michelle A. Herrmann, 2013

机译：毛细管等电聚焦免疫测定，用于研究非小细胞肺癌靶向疗法的动态癌蛋白磷酸化和药物反应

A high-throughput capillary isoelectric focusing immunoassay for fingerprinting protein sialylation

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