等压标记
化学
蛋白质组学
定量蛋白质组学
蛋白质组
色谱法
串联质量标签
肽
微流控
等压法
多路复用
无标记量化
质谱法
串联质谱法
生物分析
细胞培养中氨基酸的稳定同位素标记
组合化学
纳米技术
生物化学
蛋白质质谱法
计算机科学
电信
热力学
基因
物理
材料科学
作者
Jan Leipert,Max K Steinbach,Andreas Tholey
标识
DOI:10.1021/acs.analchem.1c01205
摘要
Digital microfluidics (DMF) is a technology suitable for bioanalytical applications requiring miniaturized, automated, and multiplexed liquid handling. Its use in LC-MS-based proteomics, however, has so far been limited to qualitative proteome analyses. This is mainly due to the need for detergents that enable facile, reproducible droplet movement, which are compatible with organic solvents commonly used in targeted chemical modifications of peptides. Aiming to implement isobaric peptide labeling, a widely applied technique allowing multiplexed quantitative proteome studies, on DMF devices, we tested different commercially available detergents. We identified the maltoside-based detergent 3-dodecyloxypropyl-1-β-d-maltopyranoside (DDOPM) to enable facile droplet movement and show micelle formation even in the presence of organic solvent, which is necessary for isobaric tandem mass tag (TMT) labeling. The detergent is fully compatible with reversed phase LC-MS, not interfering with peptide identification. Tryptic digestion in the presence of DDOPM was more efficient than without detergent, resulting in more protein identifications. Using this detergent, we report the first on-DMF chip isobaric labeling strategy, with TMT-labeling efficiency comparable to conventional protocols. The newly developed labeling protocol was evaluated in the multiplexed analyses of a protein standard digest spiked into 25 cells. Finally, using only 75 cells per biological replicate, we were able to identify 39 proteins being differentially abundant after treatment of Jurkat T cells with the anticancer drug doxorubicin. In summary, we demonstrate an important step toward multiplexed quantitative proteomics on DMF, which, in combination with larger chip arrays and optimized hardware, could enable high throughput low cell number proteomics.
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