Quigley A, Williams D R
Surfaces and Particle Engineering Laboratory, Department of Chemical Engineering, Imperial College London, SW7 2BY, UK.
Surfaces and Particle Engineering Laboratory, Department of Chemical Engineering, Imperial College London, SW7 2BY, UK.
J Chromatogr A. 2016 Aug 12;1459:47-56. doi: 10.1016/j.chroma.2016.06.048. Epub 2016 Jun 16.
Self-interaction chromatography (SIC) has established itself as an important experimental technique for the measurement of the second osmotic virial coefficients B22. B22 data are critical for understanding a range of protein solution phenomena, particularly aggregation and crystallisation. A key limitation to the more extensive use of SIC is the need to develop a method for immobilising each specific protein of interest onto a chromatographic support. This requirement is both a time and protein consuming constraint, which means that SIC cannot be used as a high throughput method for screening a wide range of proteins and their variants. Here an experimental framework is presented for estimating B22 values using Similar Interaction Chromatography (SimIC). This work uses experimental B23 and B32 data for lysozyme, lactoferrin, catalase and concanavalin A to reliably estimate B22 using arithmetic mean field approximations and is demonstrated to give good agreement with SIC measurements of B22 for the same proteins. SimIC could form the basis of a rapid protein variant screening methods to assess the developability of protein therapeutic candidates for industrial and academic researchers with respect to aggregation behaviour by eluting target proteins through a series of well-characterised protein immobilized reference columns.
自相互作用色谱法（SIC）已成为测量第二维里系数B22的一项重要实验技术。B22数据对于理解一系列蛋白质溶液现象，尤其是聚集和结晶现象至关重要。SIC更广泛应用的一个关键限制是需要开发一种将每种特定目标蛋白质固定到色谱支持物上的方法。这一要求既耗时又消耗蛋白质，这意味着SIC不能用作筛选多种蛋白质及其变体的高通量方法。本文提出了一个使用相似相互作用色谱法（SimIC）估算B22值的实验框架。这项工作利用溶菌酶、乳铁蛋白、过氧化氢酶和伴刀豆球蛋白A的实验B23和B32数据，通过算术平均场近似可靠地估算B22，并证明与相同蛋白质的SIC测量B22结果具有良好的一致性。对于工业和学术研究人员而言，SimIC可以构成一种快速蛋白质变体筛选方法的基础，通过一系列特征明确的固定有蛋白质的参考柱洗脱目标蛋白质，从而评估蛋白质治疗候选物在聚集行为方面的可开发性。
