Koichi Tanaka Laboratory of Advanced Science and Technology, Shimadzu Corporation, 1, Nishinokyo-Kuwabara-cho, Nakagyo-ku, Kyoto 604-8511, Japan.
Anal Chem. 2014 Mar 4;86(5):2549-58. doi: 10.1021/ac4037069. Epub 2014 Feb 18.
Nanoflow liquid chromatography (nano-LC) is an essential technique for highly sensitive analysis of complex biological samples, and matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) is advantageous for rapid identification of proteins and in-depth analysis of post-translational modifications (PTMs). A combination of nano-LC and MALDI-MS (nano-LC/MALDI-MS) is useful for highly sensitive and detailed analysis in life sciences. However, the existing system does not fully utilize the advantages of each technique, especially in the interface of eluate transfer from nano-LC to a MALDI plate. To effectively combine nano-LC with MALDI-MS, we integrated a nano-LC column and a deposition probe for the first time (column probe) and incorporated it into a nano-LC/MALDI-MS system. Spotting nanoliter eluate droplets directly from the column onto the MALDI plate prevents postcolumn diffusion and preserves the chromatographic resolution. A DHB prespotted plate was prepared to suit the fabricated column probe to concentrate the droplets of nano-LC eluate. The performance of the advanced nano-LC/MALDI-MS system was substantiated by analyzing protein digests. When the system was coupled with multidimensional liquid chromatography (MDLC), trace amounts of glycopeptides that spiked into complex samples were successfully detected. Thus, a nano-LC/MALDI-MS direct-spotting system that eliminates postcolumn diffusion was constructed, and the efficacy of the system was demonstrated through highly sensitive analysis of the protein digests or spiked glycopeptides.
纳升液相色谱（nano-LC）是分析复杂生物样品的高度敏感分析的重要技术，基质辅助激光解吸/电离质谱（MALDI-MS）有利于快速鉴定蛋白质和深入分析翻译后修饰（PTMs）。nano-LC 和 MALDI-MS 的组合（nano-LC/MALDI-MS）对于生命科学中的高度敏感和详细分析很有用。然而，现有的系统并没有充分利用每种技术的优势，特别是在从 nano-LC 到 MALDI 板的洗脱液转移接口。为了有效地将 nano-LC 与 MALDI-MS 结合，我们首次集成了 nano-LC 柱和沉积探针（柱探针），并将其纳入 nano-LC/MALDI-MS 系统。直接从柱上点 nano-LC 洗脱液纳米级液滴到 MALDI 板上，可防止柱后扩散并保留色谱分辨率。制备了 DHB 预点样板以适应制造的柱探针，以浓缩 nano-LC 洗脱液的液滴。通过分析蛋白质消化物证实了先进的 nano-LC/MALDI-MS 系统的性能。当系统与多维液相色谱（MDLC）偶联时，成功检测到复杂样品中添加的痕量糖肽。因此，构建了一种消除柱后扩散的 nano-LC/MALDI-MS 直接点样系统，并通过对蛋白质消化物或添加的糖肽进行高度敏感分析证明了该系统的功效。
