Bruce J E, Anderson G A, Udseth H R, Smith R D
Environmental and Molecular Sciences Laboratory, Pacific Northwest National Laboratory, P.O. Box 999, Richland, Washington 99352.
Anal Chem. 1998 Feb 1;70(3):519-25. doi: 10.1021/ac9711706.
We report a new method for mass spectrometric measurements of high-molecular-weight species based on the summation of sequential Fourier transform ion cyclotron resonance (FTICR) spectra of individual multiply charged ions. This approach produces statistically useful mass spectra for large multiply charged molecular species formed by electrospray ionization and circumvents conventional limitations upon achievable resolving power and precision for high-molecular-weight species which arise due to Coulombic constraints. For very large molecules with tens to thousands of charges each, the total number of charges required to define the charge-state distribution, and thus provide accurate mass information, greatly exceeds the useful charge capacity of the FTICR cell. As trapped ion populations approach or exceed this capacity, FTICR performance degrades due to large frequency shifts, peak coalescence phenomena, and rapid loss of ion packet coherence, which effectively precludes high-resolution and precision measurements for molecules above ∼80-kDa size for a 7-T magnetic field strength. The present approach is based on the summation of many spectra having moderate populations of individual ions and relies on sensitivity sufficient for individual ion detection. While the number of trapped ions contributing to each mass spectrum may generally be insufficient to define the isotopic or charge-state distributions (and thus produce accurate information on the molecular weight distribution in a conventional fashion), the present data processing and summation approach suppresses the noise component (as well as smaller signals) that would otherwise be problematic. Importantly, this approach circumvents natural limitations for very high molecular weight species due to Coulombic interactions and thus provides a basis for much greater resolution and mass measurement accuracy than otherwise possible. This paper presents the details of this approach and its demonstration for the 66-kDa protein bovine serum albumin (where the conventional approach is also feasible) and discusses important aspects of the data manipulation.
我们报告了一种基于对单个多电荷离子的连续傅里叶变换离子回旋共振（FTICR）光谱进行累加的方法，用于对高分子量物种进行质谱测量。这种方法为通过电喷雾电离形成的大型多电荷分子物种产生了具有统计学意义的有用质谱，并规避了由于库仑约束而导致的对高分子量物种可实现的分辨率和精度的传统限制。对于每个带有数十到数千个电荷的非常大的分子，定义电荷态分布所需的总电荷数，从而提供准确的质量信息，大大超过了FTICR池的有用电荷容量。当捕获的离子群体接近或超过此容量时，由于大的频率偏移、峰合并现象以及离子包相干性的快速丧失，FTICR性能会下降，这有效地排除了对7-T磁场强度下约80-kDa以上分子进行高分辨率和高精度测量的可能性。本方法基于对许多具有适度单个离子群体的光谱进行累加，并依赖于足以进行单个离子检测的灵敏度。虽然每个质谱中贡献的捕获离子数量通常可能不足以定义同位素或电荷态分布（从而以传统方式产生关于分子量分布的准确信息），但目前的数据处理和累加方法抑制了否则会有问题的噪声成分（以及较小的信号）。重要的是，这种方法规避了由于库仑相互作用而对非常高分子量物种的自然限制，从而为比其他可能情况更高的分辨率和质量测量精度提供了基础。本文介绍了这种方法的细节及其在66-kDa蛋白质牛血清白蛋白上的演示（在该蛋白上传统方法也是可行的），并讨论了数据处理的重要方面。
