IBET, Apartado 12, P-2781-901 Oeiras, Portugal.
J Chromatogr A. 2010 Mar 26;1217(13):2032-41. doi: 10.1016/j.chroma.2010.01.068. Epub 2010 Feb 1.
Surface plasmon resonance (SPR) spectroscopy is used as a scaled-down, analytical, pseudo-chromatography tool for analyzing protein binding and elution over an ion-exchange surface under cyclic sorption conditions. A micrometric-scale adsorption surface was produced by immobilizing a typical ion exchange ligand--diethylaminoethyl (DEAE)--onto commercially available planar gold sensor chip surfaces pre-derivatized with a self-assembled monolayer of 11-mercaptoundecanoic acid with known density. An explicit mathematical formulation is provided for the deconvolution and interpretation of the SPR sensorgrams. An adsorption rate model is proposed to describe the SPR sensorgrams for bovine serum albumin, used here as model protein, when the DEAE surface is subjected to a cyclic series of binding and elution steps. Overall, we demonstrate that the adsorption rate model is capable of quantitatively describing BSA binding and elution for protein titers from dilute conditions up to overloaded conditions and a broad range of salt concentrations.
表面等离子体共振（SPR）光谱学被用作一种缩小规模的分析型拟色谱工具，用于分析在循环吸附条件下，离子交换表面上的蛋白质结合和洗脱情况。通过将典型的离子交换配体——二乙氨基乙基（DEAE）——固定在经过预衍生的具有已知密度的 11-巯基十一酸自组装单层的市售平面金传感器芯片表面上，产生了一个微米级的吸附表面。为了解卷积和解释 SPR 传感器图，提供了一个明确的数学公式。提出了一个吸附速率模型，用于描述当 DEAE 表面经历一系列结合和洗脱步骤时，牛血清白蛋白（BSA）作为模型蛋白的 SPR 传感器图。总体而言，我们证明了吸附速率模型能够定量描述 BSA 结合和洗脱，包括从稀释条件到过饱和条件以及广泛的盐浓度范围内的情况。
