Integration of Protein Processing Steps on a Droplet Microfluidics Platform for MALDI-MS Analysis
Integration of Protein Processing Steps on a Droplet Microfluidics Platform for MALDI-MS Analysis
复制标题
DOI：
10.1021/ac9029373
复制
发表时间：
2010-03-01
影响因子：
7.4
通讯作者：
Garrell, Robin L.
中科院分区：
文献类型：
--
作者：
Chatterjee, Debalina;Ytterberg, A. Jimmy;Garrell, Robin L.
已经开发了一个基于液滴的（数字）微流体平台来制备和纯化蛋白质样品，以通过基质辅助激光解吸/电离质谱法（MALDI-MS）进行测量。液滴通过依次将电势施加到其疏水介电层下方的一系列电极中，将液滴移入空气中。我们表明，可以在该平台上执行完整的蛋白质加工步骤序列，包括二硫化物还原，烷基化和酶消化，然后使用MALDI基质和MALDI-MS对样品进行共结构化。蛋白质碳酸酐酶，细胞色素C和泛素用于证明消化和消化后步骤。胰岛素，血清白蛋白和溶菌酶用于说明平台可用的蛋白质加工步骤的完整序列，据报道，Platforrn的几种功能改进，尤其是在蛋白质液滴中掺入乙腈以促进运动，并使设备构成设备的模式。表面以优化样品结晶。该方法快速，简单，可重复，并且比常规技术的蛋白质组学样品制备相比，导致试剂消耗和样品损失较低。
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 it 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 platforrn 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.