Analytical band centrifugation revisited
Analytical band centrifugation revisited
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DOI：
10.1007/s00249-018-1315-1
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发表时间：
2018-10-01
影响因子：
2
通讯作者：
Coelfen, Helmut
中科院分区：
文献类型：
--
作者：
Schneider, Cornelia M.;Coelfen, Helmut
分析带离心（ABC）是分析大分子和纳米颗粒的有力工具。虽然它提供了比沉降速度（SV）实验的几个优点，如单个组分的物理分离和进行化学反应的可能性，它的分析仍然非常有限。因此，我们研究了ABC数据的整合作为一种替代方法，因为这会产生类似于SV的数据，然后可以通过许多已建立的评估程序进行评估。我们使用两种不同的测试系统研究了这种方法，肌红蛋白作为具有显著扩散的生物聚合物，100 nm聚苯乙烯乳胶作为具有可忽略扩散的大颗粒，并发现了一些限制性问题。即，可以考虑的是通过样品的扩散而使初始边界变宽，以及扇区中的溶剂和覆盖溶液之间的动态密度梯度，其在移动通过梯度时使初始带变形，并且目前没有考虑。我们显示了这两个因素的影响的评价，并表明，它是可以计算的时间依赖性变化的溶剂密度和粘度的AUC细胞使用的集成形式的菲克第二定律。我们的结论是，考虑到动态密度梯度将打开ABC的复杂的方法的基础上分析的整个沉积边界，而不仅仅是确定的平均沉降系数。
Analytical band centrifugation (ABC) is a powerful tool for the analysis of macromolecules and nanoparticles. Although it offers several advantages over the sedimentation velocity (SV) experiment like a physical separation of the individual components and the possibility to perform chemical reactions, its analysis is still very restricted. Therefore, we investigated the integration of ABC data as an alternative approach, as this results in data similar to SV, which can then be evaluated by many established evaluation programs. We investigated this method using two different test systems, myoglobin as a biopolymer with significant diffusion and 100 nm polystyrene latex as a large particle with negligible diffusion, and found some limiting issues. These are namely, broadening of the initial boundary by diffusion of the sample, which can be taken into account and the dynamic density gradient between the solvent in the sector and the overlaid solution, which deforms the initial band upon movement through the gradient and is currently not taken into account. We show the influence these two factors have on the evaluation and show that it is possible to calculate the time-dependent change in solvent density and viscosity in the AUC cell using the integrated form of Fick's second law. We conclude that taking the dynamic density gradient into account will open ABC for the sophisticated methods based on the analysis of the whole sedimentation boundary and not just the determination of an average sedimentation coefficient.