Mass spectrometry-based proteomics using Q Exactive, a high-performance benchtop quadrupole Orbitrap mass spectrometer.
Mass spectrometry-based proteomics using Q Exactive, a high-performance benchtop quadrupole Orbitrap mass spectrometer.
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DOI：
10.1074/mcp.m111.011015
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发表时间：
2011-09
期刊：
影响因子：
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通讯作者：
Horning S
中科院分区：
文献类型：
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作者：
Michalski A;Damoc E;Hauschild JP;Lange O;Wieghaus A;Makarov A;Nagaraj N;Cox J;Mann M;Horning S
近年来，基于质谱学的蛋白质组学从高分辨率仪器的巨大进步中受益匪浅。特别是，线性离子陷阱与Orbitrap分析仪的组合已被证明是一种流行的仪器配置。作为对这种混合陷阱-陷阱仪器的补充，以及称为Exactive的独立轨道诱捕分析器，我们在这里展示了四极质量过滤器与轨道诱捕分析器的耦合。这种“Q Exactive”仪器的特点是由于S透镜的高离子电流，以及由于平行的填充和检测模式而快速的高能碰撞诱导的解离多肽碎裂。来自探测器的图像电流通过“增强型傅立叶变换”算法处理，将质谱学分辨率提高一倍。再加上几乎瞬间的分离和碎裂，对于排名前十的高能碰撞解离方法，该仪器的总循环时间达到1 S。在哺乳动物细胞裂解液的标准90分钟胰酶消化梯度中，可以识别超过2500种蛋白质-这是对以前Orbitrap质谱仪的重大改进。此外，四极Orbitrap分析仪组合可实现MS和串联MS级别的多路传输操作。这在多路单离子监测模式中得到了证明，在这种模式下，四极杆在不同的窄质量范围内快速切换，这些窄质量范围在单个复合MS谱中进行分析。同样，四极杆允许快速连续地裂解不同的前体质量，然后在Orbitrap分析仪中联合分析更高能量的碰撞解离碎片离子。强健的台式机格式的高性能，以及执行复杂多路复用扫描模式的能力，使Q Exactive成为蛋白质组学和一般分析社区的一种令人兴奋的新仪器。
Mass spectrometry-based proteomics has greatly benefitted from enormous advances in high resolution instrumentation in recent years. In particular, the combination of a linear ion trap with the Orbitrap analyzer has proven to be a popular instrument configuration. Complementing this hybrid trap-trap instrument, as well as the standalone Orbitrap analyzer termed Exactive, we here present coupling of a quadrupole mass filter to an Orbitrap analyzer. This “Q Exactive” instrument features high ion currents because of an S-lens, and fast high-energy collision-induced dissociation peptide fragmentation because of parallel filling and detection modes. The image current from the detector is processed by an “enhanced Fourier Transformation” algorithm, doubling mass spectrometric resolution. Together with almost instantaneous isolation and fragmentation, the instrument achieves overall cycle times of 1 s for a top10 higher energy collisional dissociation method. More than 2500 proteins can be identified in standard 90-min gradients of tryptic digests of mammalian cell lysate— a significant improvement over previous Orbitrap mass spectrometers. Furthermore, the quadrupole Orbitrap analyzer combination enables multiplexed operation at the MS and tandem MS levels. This is demonstrated in a multiplexed single ion monitoring mode, in which the quadrupole rapidly switches among different narrow mass ranges that are analyzed in a single composite MS spectrum. Similarly, the quadrupole allows fragmentation of different precursor masses in rapid succession, followed by joint analysis of the higher energy collisional dissociation fragment ions in the Orbitrap analyzer. High performance in a robust benchtop format together with the ability to perform complex multiplexed scan modes make the Q Exactive an exciting new instrument for the proteomics and general analytical communities.