Formulation Research, Pharma Research and Early Development F. Hoffmann-La Roche Ltd., Basel 4070, Switzerland.
J Pharm Sci. 2011 Jul;100(7):2551-62. doi: 10.1002/jps.22506. Epub 2011 Feb 3.
The purpose of this work was to compare biophysical properties of different monoclonal antibodies (mAbs). mAbs' theoretical isoelectric point (IEP) and theoretical net charge were compared with experimentally assessed values. IEP was determined by isoelectric focusing capillary electrophoresis, determination of zero electrophoretic mobility, or the minimum mutual diffusion coefficient during pH titration. Net charge was determined using electrophoretic mobility and self-diffusion coefficient. It was found that antibodies differ substantially in their biophysical properties, that is, in IEP, net charge, and zeta potential. Also, the importance of these properties was studied with respect to protein-protein interactions. This was achieved by assessing the second virial coefficient (A(2)) determined by static light scattering (SLS) and dynamic light scattering (DLS). It was found that at low ionic strength formulation conditions [20 mM histidine (His)/His-HCl buffer, pH 6.0] proteins' charge is the main driver for overall repulsive protein interactions. At high ionic strength conditions (20 mM His/His-HCl buffer, pH 6.0, + 150 mM NaCl), where counterions are shielding ionic interactions, proteins' repulsive forces were weakened, but to a different extent. Furthermore, a DLS method was developed allowing fast and easy assessment of A(2) by minimum need of material.
这项工作的目的是比较不同单克隆抗体（mAbs）的物理化学性质。将 mAbs 的理论等电点（IEP）和理论净电荷与实验评估值进行了比较。IEP 通过等电聚焦毛细管电泳、零电泳迁移率的测定或 pH 滴定过程中的最小互扩散系数来确定。净电荷通过电泳迁移率和自扩散系数来确定。结果发现，抗体在物理化学性质方面存在显著差异，即 IEP、净电荷和zeta 电位。此外，还研究了这些特性对蛋白质-蛋白质相互作用的重要性。通过评估静态光散射（SLS）和动态光散射（DLS）测定的第二维里系数（A(2)）来实现这一点。结果发现，在低离子强度制剂条件[20 mM 组氨酸（His）/His-HCl 缓冲液，pH 6.0]下，蛋白质的电荷是总体排斥蛋白质相互作用的主要驱动力。在高离子强度条件下（20 mM His/His-HCl 缓冲液，pH 6.0，+ 150 mM NaCl），抗衡离子屏蔽离子相互作用，蛋白质的排斥力减弱，但程度不同。此外，还开发了一种 DLS 方法，通过最小的材料需求，快速轻松地评估 A(2)。
