Reversible self-association increases the viscosity of a concentrated monoclonal antibody in aqueous solution
Reversible self-association increases the viscosity of a concentrated monoclonal antibody in aqueous solution
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DOI：
10.1002/jps.20347
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发表时间：
2005-09-01
影响因子：
3.8
通讯作者：
Shire, SJ
中科院分区：
文献类型：
--
作者：
Liu, J;Nguyen, MDH;Shire, SJ
本研究旨在探讨可逆蛋白自结合对浓缩单克隆抗体溶液粘度的影响。在不同的蛋白质浓度、pH值和离子强度下，用毛细管粘度计或锥板流变仪测量单克隆抗体溶液的粘度。可溶性聚集体用粒径排除色谱法、光散射法和分析超离心法测定。利用制备式超离心机和微分馏器进行沉淀平衡分析，监测高蛋白质浓度下蛋白质的自结合。所研究的一种单克隆抗体的粘度高度依赖于缓冲液和带电赋形剂的蛋白质浓度、pH和离子强度。在低离子强度条件下，该抗体在pI附近粘度最高。沉淀平衡分析表明，该抗体在高蛋白浓度下倾向于可逆自结合。在低蛋白质浓度下，这种自结合似乎很弱，用沉降速度和粒径排除色谱法无法检测到。在低粘度和高粘度样品之间形成的不可解离的可溶性聚集体的数量没有显著差异。这些结果表明，该蛋白的可逆多价自结合似乎主要由带电残基的静电相互作用介导，并导致该单克隆抗体在低离子强度条件下在溶液中具有异常高的粘度。(c) 2005 Wiley-Liss, Inc.和美国药剂师协会。
This study was conducted to investigate the effect of reversible protein self-association on the viscosity of concentrated monoclonal antibody solutions. The viscosities of the monoclonal antibody solutions were measured by either a capillary viscometer or a cone-plate rheometer at different protein concentrations, pH, and ionic strength. Soluble aggregates were determined by size exclusion chromatography, light scattering, and analytical ultracentrifugation. Self-association of protein at high protein concentration was monitored by sedimentation equilibrium analysis using a preparative ultracentrifuge and a microfractionator. The viscosity of one of the monoclonal antibodies investigated is highly dependent on protein concentration, pH, and ionic strength of buffer and charged excipients. This antibody shows the highest viscosity near its pI at low ionic strength conditions. Sedimentation equilibrium analysis suggests that this antibody tends to reversibly self-associate at high protein concentration. The self-association appears to be quite weak and is not detectable by sedimentation velocity and size exclusion chromatography at low protein concentration. There are no significant differences in the amounts of non-dissociable soluble aggregates formed between low viscosity and high viscosity samples. These results suggest that the reversible multivalent self-association of this protein appears to be mediated mainly by electrostatic interactions of charged residues and results in unusually high viscosity of this monoclonal antibody in solution at low ionic strength conditions. (c) 2005 Wiley-Liss, Inc. and the American Pharmacists Association.