Department of Chemical Engineering University of Virginia 102 Engineers' Way, Charlottesville, VA 22904-4741 USA.
J Chromatogr A. 2011 Nov 4;1218(44):8027-35. doi: 10.1016/j.chroma.2011.09.010. Epub 2011 Sep 9.
The kinetics of single and multicomponent adsorption of deamidated monoclonal antibody (mAb) charge variants is investigated using confocal laser scanning microscopy for two commercial cation exchangers, one with an open macroporous structure--UNOsphere S--and the other with charged dextran grafts--Capto S. Markedly different intraparticle concentration profiles are obtained, being very sharp for UNOsphere S, indicating pore diffusion control, but much more diffuse for Capto S, consistent with a solid or surface diffusion mechanism. For single-component adsorption, the mAb effective pore diffusivities for UNOsphere S are approximately D(e)=4.5×10(-8) and 8.3×10(-8) cm(2)/s at pH 5 and 7.5, respectively, while effective solid diffusivities for Capto S are D(s)=0.98×10(-9) and 5.0×10(-9) cm(2)/s at pH 5 and 7.5, respectively. Two-component adsorption at pH 7.5, where the deamidated variants are bound selectively also showed markedly different profiles for the two matrices. UNOsphere S showed distinct adsorption zones within the particles indicating that multicomponent transport occurs with continuous displacement of the more deamidated variant by the less deamidated one. Capto S, however, showed no spatial resolution of the variants within the particle during co-adsorption and very slow mass transfer during sequential adsorption suggesting that protein counter-diffusion is severely hindered in this material.
使用共聚焦激光扫描显微镜研究了去酰胺单克隆抗体（mAb）电荷变异体的单组分和多组分吸附的动力学，使用两种商业阳离子交换剂，一种具有开放的大孔结构-UNOsphere S-另一种具有带电葡聚糖接枝-Capto S。得到了非常不同的颗粒内浓度分布，对于 UNOsphere S 非常尖锐，表明孔扩散控制，但对于 Capto S 则更加扩散，与固体或表面扩散机制一致。对于单组分吸附，在 pH 5 和 7.5 下，UNOsphere S 的 mAb 有效孔扩散系数分别约为 D(e)=4.5×10(-8) 和 8.3×10(-8) cm(2)/s，而 Capto S 的有效固体扩散系数分别为 D(s)=0.98×10(-9) 和 5.0×10(-9) cm(2)/s 在 pH 5 和 7.5 下。在 pH 7.5 下进行的两种组分吸附，其中去酰胺变体被选择性地结合，两种基质的吸附曲线也表现出明显的不同。在颗粒内出现明显的吸附区域，表明多组分传输发生，更去酰胺变体连续取代较少去酰胺变体。然而，Capto S 在共吸附过程中，变体在颗粒内没有空间分辨率，在顺序吸附过程中传质非常缓慢，这表明在这种材料中蛋白质的反向扩散受到严重阻碍。
