Ethier Martin, Hou Weimin, Duewel Henry S, Figeys Daniel
Ottawa Institute of Systems Biology, Ottawa, Canada.
J Proteome Res. 2006 Oct;5(10):2754-9. doi: 10.1021/pr060312m.
Gel-free proteomics has emerged as a complement to conventional gel-based proteomics. Gel-free approaches focus on peptide or protein fractionation, but they do not address the efficiency of protein processing. We report the development of a microfluidic proteomic reactor that greatly simplifies the processing of complex proteomic samples by combining multiple proteomic steps. Rapid extraction and enrichment of proteins from complex proteomic samples or directly from cells are readily performed on the reactor. Furthermore, chemical and enzymatic treatments of proteins are performed in 50 nL effective volume, which results in an increased number of generated peptides. The products are compatible with mass spectrometry. We demonstrated that the proteomic reactor is at least 10 times more sensitive than current gel-free methodologies with one protein identified per 440 pg of protein lysate injected on the reactor. Furthermore, as little as 300 cells can be directly introduced on the proteomic reactor and analyzed by mass spectrometry.
无凝胶蛋白质组学已成为传统基于凝胶的蛋白质组学的补充。无凝胶方法侧重于肽或蛋白质分级分离，但它们并未涉及蛋白质处理的效率。我们报告了一种微流控蛋白质组学反应器的开发，该反应器通过结合多个蛋白质组学步骤极大地简化了复杂蛋白质组学样品的处理。从复杂蛋白质组学样品或直接从细胞中快速提取和富集蛋白质可在该反应器上轻松完成。此外，蛋白质的化学和酶处理在50 nL有效体积内进行，这导致产生的肽数量增加。产物与质谱兼容。我们证明，该蛋白质组学反应器的灵敏度比当前无凝胶方法至少高10倍，在反应器上注入每440 pg蛋白质裂解物可鉴定出一种蛋白质。此外，仅300个细胞就可直接引入蛋白质组学反应器并通过质谱分析。
