Ultrarapid desalting of protein solutions for electrospray mass spectrometry in a microchannel laminar flow device
Ultrarapid desalting of protein solutions for electrospray mass spectrometry in a microchannel laminar flow device
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DOI：
10.1021/ac050902o
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发表时间：
2005-11-01
影响因子：
7.4
通讯作者：
Konermann, L
中科院分区：
文献类型：
--
作者：
Wilson, DJ;Konermann, L
非挥发性盐对蛋白质和其他生物分析物的电喷雾（ESI）质谱的不利影响是电喷雾质谱广泛应用的主要障碍。已经设计了许多样品清理方法来促进ESI-MS分析。最近开发的微透析技术可以将脱盐时间缩短到几分钟，其瓶颈是污染物通过半透膜的扩散。这项工作引入了一种方法，可以在几十毫秒内在线脱盐大分子溶液。该装置不使用薄膜；相反，它使用双层层流几何结构，利用大分子分析物和低分子量污染物的差异扩散。为了最大限度地提高脱盐效率，流动层之间的扩散交换只允许在允许充分交换盐的情况下进行，同时产生最小的大分子交换。计算机模拟和光学研究表明，该装置可以将盐浓度降低大约1个数量级，同时保持原始蛋白质浓度的70%左右。将该方法应用于ESI-MS中盐污染蛋白溶液的在线纯化，显著提高了信噪比和绝对信号强度。然而，有效的脱盐要求盐和分析物的扩散系数相差大约1个数量级或更多。该技术有潜力促进从复杂基质中直接进行生物大分子的高通量分析。此外，它可能成为过程监测和生物系统在线动力学研究的有价值的工具。
The adverse effects of nonvolatile salts on the electrospray (ESI) mass spectra of proteins and other biological analytes are a major obstacle for a wide range of applications. Numerous sample cleanup approaches have been devised to facilitate ESI-MS analyses. Recently developed microdialysis techniques can shorten desalting times down to several minutes, the bottleneck being diffusion of the contaminant through a semipermeable membrane. This work introduces an approach that allows the on-line desalting of macromolecule solutions within tens of milliseconds. The device does not employ a membrane; instead, it uses a two-layered laminar flow geometry that exploits the differential diffusion of macromolecular analytes and low molecular weight contaminants. To maximize desalting efficiency, diffusive exchange between the flow layers is permitted only for such a time as to allow full exchange of salt, while incurring minimal macromolecule exchange. Computer simulations and optical studies show that the device can reduce the salt concentration by roughly 1 order of magnitude, while retaining similar to 70% of the original protein concentration. Application of this approach to the on-line purification of salt-contaminated protein solutions in ESI-MS results in dramatic improvements of both the signal-to-noise ratio and the absolute signal intensity. However, efficient desalting requires the diffusion coefficients of salt and analyte to differ by roughly 1 order of magnitude or more. This technique has potential to facilitate high-throughput analyses of biological macromolecules directly from complex matrixes. In addition, it may become a valuable tool for process monitoring and for on-line kinetic studies on biological systems.