Departments of Chemistry and Molecular Biosciences, The Chemistry of Life Processes Institute , The Proteomics Center of Excellence at Northwestern University , Evanston , Illinois 60208 , United States.
Thermo Fisher Scientific , San Jose , California 95134 , United States.
Anal Chem. 2019 Feb 19;91(4):2776-2783. doi: 10.1021/acs.analchem.8b04519. Epub 2019 Jan 29.
It is well-known that with Orbitrap-based Fourier-transform-mass-spectrometry (FT-MS) analysis, longer-time-domain signals are needed to better resolve species of interest. Unfortunately, increasing the signal-acquisition period comes at the expense of increasing ion decay, which lowers signal-to-noise ratios and ultimately limits resolution. This is especially problematic for intact proteins, including antibodies, which demonstrate rapid decay because of their larger collisional cross-sections, and result in more frequent collisions with background gas molecules. Provided here is a method that utilizes numerous low-ion-count spectra and single-ion processing to reconstruct a conventional m/ z spectrum. This technique has been applied to proteins varying in molecular weight from 8 to 150 kDa, with a resolving power of 677 000 achieved for transients of carbonic anhydrase (29 kDa) with a duration of only ∼250 ms. A resolution improvement ranging from 10- to 20-fold was observed for all proteins, providing isotopic resolution where none was previously present.
众所周知，基于轨道阱的傅里叶变换质谱（FT-MS）分析需要更长的时域信号，以便更好地解析感兴趣的物质。然而，增加信号采集时间会导致离子衰减增加，从而降低信噪比，最终限制分辨率。这对于完整的蛋白质（包括抗体）来说尤其成问题，因为它们的碰撞截面较大，所以衰减较快，与背景气体分子的碰撞也更频繁。本文提供了一种利用多次低离子计数谱和单离子处理来重建常规 m/z 谱的方法。该技术已应用于分子量从 8 到 150 kDa 不等的蛋白质，对于持续时间仅约 250 ms 的碳酸酐酶（29 kDa）瞬变，实现了 677 000 的分辨率。所有蛋白质的分辨率都提高了 10 到 20 倍，在以前没有同位素分辨率的情况下提供了同位素分辨率。
