A Dual Pressure Linear Ion Trap Orbitrap Instrument with Very High Sequencing Speed

Since its introduction a few years ago, the linear ion trap Orbitrap (LTQ Orbitrap) instrument has become a powerful tool in proteomics research. For high resolution mass spectrometry measurements ions are accumulated in the linear ion trap and passed on to the Orbitrap analyzer. Simultaneously with acquisition of this signal, the major peaks are isolated in turn, fragmented and recorded at high sensitivity in the linear ion trap, combining the strengths of both mass analyzer technologies. Here we describe a next generation LTQ Orbitrap system termed Velos, with significantly increased sensitivity and scan speed. This is achieved by a vacuum interface using a stacked ring radio frequency ion guide with 10-fold higher transfer efficiency in MS/MS mode and 3–5-fold in full scan spectra, by a dual pressure ion trap configuration, and by reduction of overhead times between scans. The first ion trap efficiently captures and fragments ions at relatively high pressure whereas the second ion trap realizes extremely fast scan speeds at reduced pressure. Ion injection times for MS/MS are predicted from full scans instead of performing automatic gain control scans. Together these improvements routinely enable acquisition of up to ten fragmentation spectra per second. Furthermore, an improved higher-energy collisional dissociation cell with increased ion extraction capabilities was implemented. Higher-collision energy dissociation with high mass accuracy Orbitrap readout is as sensitive as ion trap MS/MS scans in the previous generation of the instrument.Proteomics experiments typically involve the analysis of peptide mixtures obtained by the enzymatic digestion of proteomes that can be as complex as complete cell lysates (1, 2). Dynamic range of peptide abundances and the sheer number of peptides encountered in these mixtures require extremely sensitive and fast peptide detection and fragmentation (3). Although a first comprehensively identified and quantified proteome has recently been reported (4), further gains in instrumental performance are clearly needed to reduce overall measurement time, improve sequence coverage of identified proteins, and for the in-depth analysis of mammalian proteomes.Among many different instrumental formats (5), the combination of a linear ion trap (6) with a Fourier transform (FT)1 mass spectrometer has rapidly become a popular technological platform in proteomics because it combines the sensitivity, speed, and robustness of ion traps with the high resolution capabilities of FT instruments. The first implementation of this principle used an ion cyclotron resonance instrument with a 7T magnet as the high resolution device (7). Later, the Orbitrap^TM analyzer developed by Makarov was coupled to the LTQ, combining the linear ion trap with a very small and powerful analyzer (8–11).Here we describe a next generation linear ion trap-Orbitrap instrument with significant improvements in ion source transmission and with a new ion trap configuration. We show that this instrument, termed the LTQ Orbitrap Velos, is capable of much higher scan speeds compared with the current LTQ Orbitrap. Furthermore, we implemented more efficient ion extraction for the higher-energy collisional dissociation (HCD) cell (12). Due to this improvement and the 10-fold higher transmission of ions from atmosphere, high resolution and high mass accuracy MS/MS can now routinely be obtained at very high sensitivity and at scan speeds of up to 5 Hz acquisition rates. A related instrument, the LTQ-Velos, which does not contain the Orbitrap analyzer for high resolution measurements, has been described very recently (13).