College of Chemistry, University of California, Berkeley, Berkeley, California 94720-1460, United States.
California Institute for Quantitative Biosciences, University of California, Berkeley, Berkeley, California 94720-1460, United States.
Anal Chem. 2022 Aug 23;94(33):11703-11712. doi: 10.1021/acs.analchem.2c02572. Epub 2022 Aug 12.
Instrumental resolution of Fourier transform-charge detection mass spectrometry instruments with electrostatic ion trap detection of individual ions depends on the precision with which ion energy is determined. Energy can be selected using ion optic filters or from harmonic amplitude ratios (HARs) that provide Fellgett's advantage and eliminate the necessity of ion transmission loss to improve resolution. Unlike the ion energy-filtering method, the resolution of the HAR method increases with charge (improved /) and thus with mass. An analysis of the HAR method with current instrumentation indicates that higher resolution can be obtained with the HAR method than the best resolution demonstrated for instruments with energy-selective optics for ions in the low MDa range and above. However, this gain is typically unrealized because the resolution obtainable with molecular systems in this mass range is limited by sample heterogeneity. This phenomenon is illustrated with both tobacco mosaic virus (0.6-2.7 MDa) and AAV9 (3.7-4.7 MDa) samples where mass spectral resolution is limited by the sample, including salt adducts, and not by instrument resolution. Nevertheless, the ratio of full to empty AAV9 capsids and the included genome mass can be accurately obtained in a few minutes from 1× PBS buffer solution and an elution buffer containing 300+ mM nonvolatile content despite extensive adduction and lower resolution. Empty and full capsids adduct similarly indicating that salts encrust the complexes during late stages of droplet evaporation and that mass shifts can be calibrated in order to obtain accurate analyte masses even from highly salty solutions.
傅里叶变换-电荷检测质谱仪器的仪器分辨率取决于离子能量确定的精度。可以使用离子光学滤波器或从提供 Fellgett 优势并消除离子传输损失以提高分辨率的必要性的谐波幅度比 (HAR) 来选择能量。与离子能量过滤方法不同，HAR 方法的分辨率随电荷（改善 /）和质量增加而增加。对当前仪器的 HAR 方法进行的分析表明，与具有能量选择光学器件的仪器所能达到的最佳分辨率相比，HAR 方法可以获得更高的分辨率，这些仪器适用于低 MDa 及以上范围内的离子。然而，通常无法实现这种增益，因为在这个质量范围内，分子系统可获得的分辨率受到样品异质性的限制。这一现象通过烟草花叶病毒（0.6-2.7 MDa）和 AAV9（3.7-4.7 MDa）的样品来说明，其中质谱分辨率受到样品的限制，包括盐加合物，而不是仪器分辨率。尽管存在广泛的加合物和较低的分辨率，但从 1×PBS 缓冲溶液和含有 300+ mM 非挥发性物质的洗脱缓冲液中，仅几分钟即可准确获得 AAV9 空壳和满壳的比例以及包含的基因组质量。空壳和满壳加合物类似，表明盐在液滴蒸发的后期阶段包裹在复合物上，并且可以对质量偏移进行校准，以便即使从高盐溶液中也可以获得准确的分析物质量。
