Department of Chemistry and Biochemistry, University of California-Los Angeles, Los Angeles, California 90095-1569, USA.
Anal Chem. 2010 Mar 1;82(5):2095-101. doi: 10.1021/ac9029373.
A droplet-based (digital) microfluidics platform has been developed to prepare and purify protein samples for measurement by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). Liquid droplets are moved in air by sequentially applying an electric potential to an array of electrodes patterned beneath a hydrophobic dielectric layer. We show that a complete integrated sequence of protein processing steps can be performed on this platform, including disulfide reduction, alkylation, and enzymatic digestion, followed by cocrystallization with a MALDI matrix and analysis of the sample in situ by MALDI-MS. Proteins carbonic anhydrase, cytochrome c, and ubiquitin were used to demonstrate the digestion and postdigestion steps; insulin, serum albumin, and lysozyme were used to illustrate the complete sequence of protein processing steps available with the platform. Several functional improvements in the platform are reported, notably, the incorporation of acetonitrile in the protein droplets to facilitate movement, and patterning the device surfaces to optimize sample crystallization. The method is fast, simple, repeatable, and results in lower reagent consumption and sample loss than conventional techniques for proteomics sample preparation.
已经开发出一种基于液滴（数字）微流控平台，用于通过基质辅助激光解吸/电离质谱（MALDI-MS）来准备和纯化蛋白质样品。通过顺序向疏水介电层下方的电极阵列施加电势，可以在空气中移动液滴。我们表明，可以在该平台上执行完整的蛋白质处理步骤的集成序列，包括二硫键还原、烷基化和酶消化，然后与 MALDI 基质共结晶，并通过 MALDI-MS 原位分析样品。使用碳酸酐酶、细胞色素 c 和泛素来证明消化和消化后步骤；使用胰岛素、血清白蛋白和溶菌酶来说明该平台可用的完整蛋白质处理步骤序列。报道了该平台的几个功能改进，特别是在蛋白质液滴中加入乙腈以促进移动，以及对设备表面进行图案化以优化样品结晶。该方法快速、简单、可重复，并且与传统的蛋白质组学样品制备技术相比，消耗的试剂和样品损失更少。
