Hybrid mass spectrometer and data acquisition method

Abstract

To provide a method of operating a dual analyzer mass spectrometer to obtain MS1 scans / MS2 scans of positive and negative ions from a sample.SOLUTION: The method comprises: operating an ion source and a processing region at a first polarity; performing MS1 or MS2 scan(s) by a first mass analyzer at the first polarity; switching the polarity of the first mass analyzer to a second polarity; and performing MS1 or MS2 scans by a second mass analyzer at the first polarity, where at least part of the MS1 or MS2 scans performed by the second mass analyzer are performed during a deadtime in which the first mass analyzer is switching the polarity.SELECTED DRAWING: Figure 2

Description

Technical Field

[0001] The present invention relates to a method of operating a dual analyzer mass spectrometer to obtain MS1 and MS2 scans of ions from a sample, for example, when eluting from a chromatographic system. Methods are provided for obtaining scans of positive and negative ions. An apparatus for carrying out the method is also provided.

Background Art

[0002] Mass spectrometry is often a long-established technique for the identification and quantification of a wide range of biological and non-biological materials, including complex mixtures of large organic molecules.

[0003] Proteins containing a large number of amino acids typically have a fairly large molecular weight. Therefore, accurate identification and quantification of proteins by direct mass spectrometry measurements are difficult. It is well known to perform fragmentation of ionized sample material, considered to be precursor ions. Various fragmentation techniques are well known that can result in the generation of different fragment ions from precursor ions.

[0004] To determine the molecular structure of sample molecules, a mass spectrometer may first be used to mass analyze all sample ions (precursor ions) within a selected mass-to-charge ratio (m / z) range. Such a scan often means an MS1 scan. Second, the sample ions may be fragmented and mass analyzed. In this second step, sample ions from a narrow window of the mass charge range may be selected, fragmented, and the resulting fragments may be mass analyzed. After fragmentation, the ions may similarly have a wide mass-to-charge ratio range, so the mass analyzer itself may still scan a wide range of mass-to-charge ratios, as in an MS1 scan. The scan of the fragmented ions is often shown as an MS2 scan. Since the narrow window of the m / z range selected for each MS2 may be smaller than the window of the precursor ion scan, multiple MS2 scans are generally performed. Multiple MS2 scans may be used to cover the desired range of sample ion m / z.

[0005] Two methods for identifying the range covered by MS2 scanning are Data Independent Analysis or acquisition (DIA) and Data Dependent Analysis or acquisition (DDA). The Data Independent Analysis / Acquisition method (DIA) may be used to determine what is present in a sample of potentially unknown identity. In this method, first, a mass spectrometer is used to mass analyze all sample ions (precursor ions) within a selected mass-to-charge ratio (m / z) range to obtain an MS1 scan. In the MS2 scan, the target mass range is simply segmented, and an MS2 spectrum is obtained for each segment. In the case of DIA, the MS1 scan may be optional because the selection window parameters themselves for determining which ions to select and which ions to perform an MS2 scan on carry information about the range of possible sample ions within the window. The Data Dependent Analysis / Acquisition method (DDA) differs from DIA in that DDA is more suitable for confirming the presence of one or more species in a sample and is used when there is substantial knowledge of the species and the structure of the sample. Thus, this knowledge of the species and structure can be used to identify a fixed or limited number of expected precursor ions and select and analyze them. Fragmentation and MS2 spectra may be performed based on an “inclusion list” of the species that are candidates to be discovered. Additionally or alternatively, fragmentation and MS2 spectra may be set based on the species determined by the MS1 scan, for example, based on the intensity ranking of the abundance of the species determined by the MS1 scan.

[0006] Most mass spectrometry research is performed through cation detection, but many analytes, such as acidic peptides and all classes of lipids, are susceptible to anionization and detection. Even up to the time scale of analyte introduction into the mass spectrometer, which can be <500 ms, preferably <50 ms, from liquid / gas chromatography separation, it is common for commercially available mass spectrometry instruments to be switchable between cation mode and anion mode, and it is possible to acquire multiple spectra over a chromatographic peak that can have a duration of about 1 second.

[0007] Triple quadrupole mass spectrometry instruments capable of performing MS1 scans and MS2 scans may require, for example, a polarity switching time of up to 5 ms, which is governed by the high voltage switching time for the electrospray ion source and detector conversion dynode (the electron multiplier section of the photomultiplier tube, dynode), with additional delays for electrospray plume stabilization and ion transport. A 5 ms polarity switching time is probably equivalent to the loss of only a few spectra in a triple quadrupole mass spectrometer, and thus experiments incorporating polarity switching may be performed without compromising instrument operation beyond the basic compromise of splitting the acquisition time between the two polarities. However, triple quadrupole mass spectrometers have a high polarity switching speed but relatively low accuracy and poor resolution compared to mass spectrometers incorporating high resolution accurate-mass (HRAM) analyzers. Next, HRAM analyzers have longer polarity switching times. Therefore, it is desirable to provide methods and apparatus that use HRAM analyzers but are not limited by their switching speeds. For example, slow switching speeds will result in a dead time where data is not collected and the presence of species may be missed.

[0008] Fast polarity switching in high-resolution accurate mass (HRAM) analysers such as time-of-flight analysers and orbital trapping mass analysers is difficult because these analysers typically require ppm-stable high-voltage potentials, which are often strongly filtered and involve corresponding long delays to charge capacitances to stabilise the voltage output. The mass analyser cannot be used during the switching period, which can last for seconds or minutes. Some commercially available HRAM instruments support pulsed polarity switching within a single experiment, but notable exceptions are modern orbital trapping mass analyser instruments such as the Orbitrap™ Exploris™ series (manufactured by Thermo Fisher Scientific), which can switch polarity within 500 ms. As described above, for chromatographic elution times of about 1 s, this is a troublesome dead time for the instrument. For this reason, pulsed polarity switching is rarely used within a single experiment.

[0009] Orbitrap™ Exploris™ instruments achieve relatively fast polarity switching by maintaining both positive and negative stable HV supplies to their critical centre poles and switching between them via high-voltage transistor switches. This approach allows for the number of components and the capacitance between the switches and the electrodes to be limited. The disadvantage is that the number of components is effectively doubled compared to a system that switches the polarity of an unregulated HV source. Therefore, this approach is expensive and takes up space.

[0010] Other efforts have been made in the prior art to provide a high-resolution, high-accuracy mass spectrometer that can operate to detect both cations and anions from a sample.

[0011] One characteristic used by Furutani in U.S. Patent No. 7,170,052(B2) is that the force acting on the ions is reversed with respect to the opposite polarity of the ions. U.S. Patent No. 7,170,052(B2) utilizes this characteristic to simultaneously ionize and analyze both ion polarities using two analyzers within a hybrid device, with one operating at each polarity. The device includes a dual orthogonal time-of-flight (ToF) analyzer, and the positive and negative ions generated by the ion source are separated by their opposite directions of ion mobility for different polarities. The separated positive and negative ions are analyzed by their respective polarity analyzers. Since polarity switching is avoided, that problem is also avoided. A similar device is described by Wang in U.S. Patent No. 7,649,170(B2).

[0012] Hybrid devices incorporating multiple analyzers have the general advantage that the strength of one analyzer can compensate for the weakness of the other. In Furutani, since two analyzers of the same type are used, such an advantage is not seen. More often, the analyzers are of different types. For example, the Orbitrap(™) Exploris(™) series (manufactured by Thermo Fisher Scientific) combines an ion trap mass analyzer and an orbitrap mass analyzer.

[0013] U.S. Patent No. 10,699,888 (B2) by Giannakopulos describes an instrument that combines an orbitrap mass spectrometer and a multi-reflection time-of-flight (MR-ToF) analyzer, equipped with a quadrupole mass filter and a collision cell for tandem mass spectrometry. This document describes generating a full mass (MS1) scan using a low-speed, high-precision orbitrap mass spectrometer while the high-speed, high-sensitivity MR-ToF simultaneously provides a fragment (MS2) spectrum. The MR-ToF analyzer is of the opposing mirror type as described by Grinfeld in U.S. Patent No. 9,136,101 (B2), which is less affected by high-speed polarity switching than an orbitrap mass spectrometer. The mirror electrode structure requires four stable high voltages and still requires a voltage higher than 5 KV of the central electrode of the orbitrap mass spectrometer.

[0014] Dual analyzer HRAM mass spectrometers are well-known, but none of them provide a solution to overcome the significant dead time of the analyzer's polarity switching when MS1 scans and / or MS2 scans are required in both polarities.

[0015] High-speed switching power supplies, such as those described for orbitrap mass spectrometer instruments, are relatively complex and expensive. Analyzers with multiple stable HV power supplies, such as MR-ToF, become large in size and complexity and are further exacerbated by the higher voltages that require even more expensive components. SUMMARY OF THE INVENTION

[0016] The present invention relates to a method of operating a dual analyzer mass spectrometer that avoids long dead times when ions or ion beams are not used because the mass spectrometer is switching polarities.

[0017] In the first embodiment, in order to avoid the dead time while the mass spectrometer(s) is switching polarities, the timing of ion acquisition and mass spectrometer switching may be offset from one analyzer to the other. In this regard, the polarity switching time of one mass spectrometer may be parallel to the ion acquisition by the other mass spectrometer and the performance of mass scanning. The period during which the ion source and the ion processing region are switching polarities may also be efficiently used, that is, by acquiring ions immediately before the switching of one of the mass spectrometers such that the mass spectrometer performs a mass scan during the dead time of the ion source and the ion processing region.

[0018] In the second embodiment, the polarity of the mass spectrometer is not changed such that one analyzer performs a mass scan with a first polarity and the other analyzer performs a mass scan with a second polarity opposite to the first polarity. The ion source and the ion processing region have their polarities switched and direct the ions to one analyzer and then to the other analyzer, respectively, according to the ion polarity. Thereby, the long polarity switching time of the mass spectrometer is avoided. Again, the period during which the ion source and the ion processing region are switching polarities is efficiently used, that is, by acquiring ions immediately before the switching of one of the mass spectrometers such that the mass spectrometer performs a mass scan during the dead time of the ion source and the ion processing region switching.

[0019] Accordingly, a first embodiment of the present invention provides a method of operating a dual analyzer mass spectrometer to obtain MS1 and MS2 scans of cations and anions from a sample. The method includes ionizing the sample in an ion source and an ion processing region of the mass spectrometer to generate a plurality of ions in an ion source and an ion processing region operating in a first polarity, directing a first ion packet of the plurality of ions to a first mass analyzer, performing a first scan sequence including at least one MS1 scan or at least one MS2 scan of the ions in the first packet in the first polarity by the first mass analyzer, switching the polarity of the first mass analyzer to a second polarity after performing at least one MS1 scan or at least one MS2 scan, directing one or more second packets of ions of the plurality of ions to a second mass analyzer, and performing a second scan sequence including at least one MS1 scan or at least one MS2 scan of the ions in the one or more second packets in the first polarity by the second mass analyzer, wherein at least a portion of at least one MS1 scan or at least a portion of at least one MS2 scan performed by the second mass analyzer is performed during a first dead time in which the first mass analyzer is switching polarities.

[0020] The terms "operating in a first polarity" or "operating in a second polarity" mean that the respective ion source, ion guiding / processing region, first mass analyzer, and / or second mass analyzer are configured to operate for the analysis of ions in the respective polarities. The first polarity may be a positive or negative polarity, and the second polarity will be opposite to the first polarity.

[0021] After ionizing the ions, the ions may be accumulated or collected in an ion trap or the like over a certain period to generate a packet of ions. The dead time is a period during which the first mass spectrometer or the second mass spectrometer cannot perform an accurate mass scan due to polarity switching, or a period during which the ion source and the ion processing region are not ionizing the sample due to polarity switching. For example, the first dead time may be the case where the first mass spectrometer is switching polarities.

[0022] The method may further include switching the polarities of the ion source and the ion processing region of the mass spectrometer to a second polarity while the second mass spectrometer performs at least one MS1 scan or at least one MS2 scan of the ions in one or more second packets in the first polarity. The method may further include repeating the step of directing the packet of ions and the step of performing scans with the first mass spectrometer and the second mass spectrometer in the second polarity.

[0023] The last MS1 scan or MS2 scan of the second scan sequence, performed by the second mass spectrometer in the first polarity, may be at least partially performed during the second dead time when the ion source and the ion processing region are switching polarities to the second polarity.

[0024] The method includes ionizing a sample in an ion source and an ion processing region operating in a second polarity to generate a further plurality of ions after switching the polarities of the ion source and the ion processing region to the second polarity, directing a third ion packet to the first mass spectrometer, and performing a third scan sequence including at least one MS1 scan or at least one MS2 scan of the ions in the third packet by the first mass spectrometer operating in the second polarity.

[0025] This method further includes, after performing at least one MS1 scan or at least one MS2 scan of ions in one or more second packets in a first polarity by a second mass analyzer, and at least one MS1 scan or at least one MS2 scan of ions in one or more second packets during a first dead time when the first mass analyzer is switching polarities, switching the polarity of the second mass analyzer to a second polarity; and directing one or more fourth packets of ions to the second mass analyzer and performing a fourth scan sequence including at least one MS1 scan or at least one MS2 scan of ions in one or more fourth packets in the second polarity by the second mass analyzer. A method of performing at least one MS1 scan or at least one MS2 scan of ions in a third packet by a first mass analyzer operating in the second polarity may be performed at least partially during a third dead time when the second mass analyzer is switching its polarity to the second polarity.

[0026] A method of performing at least one MS1 scan or at least one MS2 scan of ions in a first packet by a first mass analyzer operating in a first polarity may be performed at least partially during a fourth dead time when the second mass analyzer is switching its polarity to the first polarity. The fourth dead time may alternatively be considered the zero dead time since it may occur when the first ion packet is being sent to the first mass analyzer and also after the fourth ion packet has been sent to the second mass analyzer.

[0027] A dead time may be considered to occur when (i) the first analyzer is switching polarities, (ii) the second analyzer is switching polarities, or (iii) the ion source and ion processing region are switching polarities. Since the three dead times may be considered to occur for each polarity of operation, a method cycle including four scan sequences (such as MS1 and MS2 scans across both polarities) may be considered to include six dead times.

[0028] The third scanning sequence performed by the first mass analyzer in the second polarity may include an MS1 scan. The method directs a third ion packet to the first mass analyzer and, after starting the third scanning sequence, directs one or more fourth ion packets to the second mass analyzer without switching the polarity of the second mass analyzer, and performs one or more MS2 scans of the ions in one or more fourth ion packets in the first polarity by the second mass analyzer, with at least a portion of the one or more MS2 scans being performed while the first mass analyzer is performing an MS1 scan.

[0029] The method further includes switching the polarity of the ion source and the ion processing region to the second polarity after performing an MS1 scan in the second polarity by the first mass analyzer during the third scanning sequence, and after switching the polarity of the ion source and the ion processing region to the second polarity, directing one or more fifth packets of ions to the first mass analyzer and performing a fifth scanning sequence including one or more MS2 scans of the ions in one or more fifth packets in the second polarity by the first mass analyzer.

[0030] The second mass analyzer may not have its polarity switched and may perform MS2 scans only in the first polarity. Thus, the first mass analyzer may perform other scans, namely, MS1 and MS2 scans in the second polarity, and MS1 scans in the first polarity.

[0031] The method may be performed within a period based on the width of the chromatographic peak of the sample when the sample elutes from the chromatographic system. Generally, each cycle (e.g., corresponding to the combined duration of the first and second scanning sequences) should be similar to or less than the chromatographic peak width to capture each peak in both the positive and negative modes.

[0032] The first scanning sequence may include one or more MS1 scans, and the second scanning sequence may include one or more MS2 scans. Alternatively, the first scanning sequence may include one or more MS2 scans, and the second scanning sequence may include one or more MS1 scans. When MS2 scans are performed in the scanning sequence, the scanning sequence may include performing a plurality of MS2 scans respectively on a series of ion packets sequentially directed to each analyzer.

[0033] The first mass analyzer may be an orbitrap mass analyzer, the second mass analyzer may be an ion trap mass analyzer or a time-of-flight mass analyzer (e.g., MR-ToF), or vice versa.

[0034] The dual-analyzer mass spectrometer may include one or more ion traps such as a curved ion trap or a C trap. The method may include aggregating or collecting ions ionized by an ion source and an ion processing region by at least one of the one or more ion traps to form an ion packet, and directing the ion packet to the first mass analyzer and / or the second mass analyzer by at least one ion trap. In some embodiments, one mass analyzer may aggregate ions to form a packet and selectively direct the ion packet to the first and second mass analyzers. In other embodiments, each analyzer may have its own ion trap, each may aggregate ions to form a packet, and each may control the injection of the ion packet into its respective analyzer.

[0035] The first mass analyzer and / or the second mass analyzer require a period of at least 50 ms, at least 100 ms, at least 200 ms, or at least 500 ms to switch polarities and resume mass spectrometry.

[0036] The mass analyzer is preferably a high-resolution accurate mass (HRAM) analyzer. The high-resolution accurate mass (HRAM) analyzer preferably has a resolution of about 10,000 or more and preferably has a mass accuracy better than about 10 ppm.

[0037] The method further comprises further ionizing the sample in a first polarity to generate additional ions, directing additional first ion packets of the ions to a first mass analyzer, and performing a fifth scan sequence including at least one MS1 scan or at least one MS2 scan of the ions in the additional first packet in the first polarity by the first mass analyzer, switching the polarity of the first mass analyzer to a second polarity after performing at least one MS1 scan or at least one MS2 scan, directing one or more additional second ion packets of the ions to a second mass analyzer, and performing a sixth scan sequence including at least one MS1 scan or at least one MS2 scan of the ions in the one or more additional second packets in the first polarity by the second mass analyzer, and at least a part of at least one MS1 scan or at least a part of at least one MS2 scan performed by the second mass analyzer is performed during an additional first dead time in which the first mass analyzer is switching polarities.

[0038] The second embodiment of the present invention provides a method of operating a dual mass spectrometer to obtain MS1 and MS2 scans of cations and anions from a sample. The first mass spectrometer may operate in a first polarity, and the second mass spectrometer may operate in a second polarity opposite to the first polarity. In this embodiment, the polarity of the mass spectrometer may be maintained constant without switching or reversing the polarity. The method includes ionizing a sample in an ion source and an ion processing region of the mass spectrometer to generate a plurality of ions in the ion source and ion processing region operating in the first polarity, directing one or more first packets of ions to the first mass spectrometer, and causing the first mass spectrometer to perform at least one MS1 scan and / or at least one MS2 scan of the ions in the one or more first packets in the first polarity. After the first mass spectrometer starts at least one MS1 scan and / or at least one MS2 scan of one or more first ion packets, switching the polarity of the ion source and the ion processing region to a second polarity opposite to the first polarity, and after switching the polarity of the ion source and the ion processing region to the second polarity, directing one or more second ion packets to the second mass spectrometer and causing the second mass spectrometer to perform at least one MS1 scan and / or at least one MS2 scan of the ions in the second packet in the second polarity.

[0039] The first mass spectrometer may perform MS1 and MS2 scans in the first polarity, and the second mass spectrometer may perform MS1 and MS2 scans in the second polarity.

[0040] At least a part of at least one MS1 scan and / or at least a part of at least one MS2 scan performed by the first mass spectrometer may be performed during a dead time when the polarity of the ion source and the ion processing region is switched to the second polarity.

[0041] The period forming the MS1 scan for the first mass spectrometer and / or the second mass spectrometer may be longer than the period for performing the MS2 scan.

[0042] One or more second ion packets may include a plurality of second ion packets, and while the first mass analyzer is performing an MS1 scan and after the ion source and the ion processing region have switched polarity to a second polarity, the method includes performing, by a second mass analyzer, one or more MS1 scans and one or more MS2 scans in the second polarity on the plurality of second ion packets; switching the polarity of the ion source and the ion processing region back to the first polarity while the second mass analyzer is performing the last scan of a series of scans on the plurality of second ion packets; and directing one or more third ion packets to the first mass analyzer and performing one or more MS2 scans after switching the ion source and the ion processing region back to the first polarity.

[0043] While the first mass analyzer is performing an MS1 scan in a first polarity, the second mass analyzer may perform MS1 and MS2 scans in a second polarity.

[0044] One or more first packets of ions may include a first ion packet, and the first mass analyzer may perform an MS1 scan of the ions within the first ion packet in a first polarity, and after switching the polarity of the ion source and the ion processing region to a second polarity and directing one or more second ion packets to a second analyzer, the second analyzer may perform one or more MS2 scans on the one or more second ion packets in the second polarity.

[0045] This method may further include, after performing one or more MS2 scans on a second ion packet by a second analyzer, directing a third ion packet to the second analyzer and performing an MS1 scan of the third ion packet by the second analyzer with a second polarity; after directing the third ion packet to the second mass analyzer and performing an MS1 scan, while the second mass analyzer is performing the MS1 scan of the third ion packet, switching the polarities of the ion source and the ion processing region to a second polarity opposite to the first polarity; and after switching the polarities of the ion source and the ion processing region to the second polarity, directing one or more fourth ion packets to the first mass analyzer and performing one or more MS2 scans of the ions in the one or more fourth ion packets by the first mass analyzer with the first polarity.

[0046] This method may be performed within a period based on the width of the chromatographic peak of the sample when the sample elutes from the chromatographic system. Generally, each cycle (e.g., corresponding to the combined duration of the first scan sequence and the second scan sequence) should be similar to or less than the chromatographic peak width to capture each peak in the positive and negative modes.

[0047] The MS2 scan for one of the first mass analyzer and the second mass analyzer may be performed (temporally) in parallel with the MS1 scan performed on the other of the first mass analyzer and the second mass analyzer.

[0048] The first mass analyzer may be an orbitrap mass analyzer, the second mass analyzer may be a time-of-flight mass analyzer (e.g., MR-ToF) or an orbitrap mass analyzer, or vice versa.

[0049] The dual analyzer mass spectrometer may include one or more ion traps such as a curved ion trap or a C trap. The method may include aggregating or collecting ions ionized by an ion source and an ion processing region by at least one of the one or more ion traps to form an ion packet, and directing the ion packet by at least one of the ion traps to a first mass spectrometer and / or a second mass spectrometer. In some embodiments, one mass spectrometer may aggregate ions to form a packet and selectively direct the ion packet to a first mass spectrometer and a second mass spectrometer. In other embodiments, each analyzer may have its own ion trap, aggregate ions to form a packet respectively, and control the injection of the ion packet into each analyzer.

[0050] The first mass spectrometer and / or the second mass spectrometer may require a period of at least 50 ms, at least 100 ms, at least 200 ms, or at least 500 ms to switch polarities and resume mass spectrometry.

[0051] The mass spectrometer may be a high-resolution accurate mass (HRAM) analyzer. The high-resolution accurate mass (HRAM) analyzer preferably has a resolution of about 10,000 or more and preferably has a mass accuracy of less than about 10 ppm.

[0052] The present invention further provides a mass spectrometer comprising an ion source and an ion processing region including an ionization source for generating a plurality of precursor ions from sample molecules and a mass filter, one or more ion traps configured to aggregate / collect the ions ionized by the ion source and the ion processing region to form an ion packet and selectively direct the ion packet to a first mass analyzer and / or a second mass analyzer, a first mass analyzer, a fragmentation device, a second mass analyzer, and a controller configured to cause the mass spectrometer to perform any of the methods described herein. The present invention may further provide a system comprising a mass spectrometer and a chromatography system configured to separate sample molecules, the chromatography system providing the separated molecules to the ionization source.

[0053] The present invention provides a computer program comprising computer program instructions that, when executed on a computer or controller configured to control a mass spectrometer, cause the mass spectrometer to perform any of the steps of the methods described herein.

[0054] In this specification, two analyzers of the mass spectrometer have been described as including an orbitrap mass analyzer together with an ion trap, a ToF analyzer, or a second orbitrap mass analyzer, but other combinations of analyzers are possible. For example, the other mass analyzers may be FTICR (Fourier Transform Ion Cyclotron Resonance), quadrupole, sector, or electrostatic trap. The first analyzer and the second analyzer may include the following alternative combinations: FTICR and ToF, or electrostatic trap and ToF.

[0055] In this specification, the term "mass" may be used to refer to the mass-to-charge ratio, m / z. It should be understood that the resolution of the mass analyzer refers to the resolution of the mass analyzer determined at a mass-to-charge ratio of 200, unless otherwise specified.

[0056] Hereinafter, embodiments of the present invention and aspects of the prior art will be described with reference to the accompanying drawings.

Brief Description of the Drawings

[0057]

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DETAILED DESCRIPTION OF THE INVENTION

[0058] FIG. 1 shows a schematic configuration of a mass spectrometer 1 suitable for implementing the method according to an embodiment of the present invention. The mass spectrometer is a tandem mass spectrometer and includes an ion source 11 such as an electrospray ionization (ESI) source, and an ion processing region 12 that may include various electrodes for purifying and filtering an ion beam and generating a field for selecting a desired ion m / z range. In the configuration shown in FIG. 1, the first analyzer 13 is shown, for example, crossing the axis of the ion beam in the ion processing region 12, away from the direction of the axis of the ion beam. The second analyzer 14 is shown as terminating the direction of the ion beam in the ion focusing region 12. Other arrangements of the tandem mass spectrometer may be provided and will be described later in this specification.

[0059] The sample to be analyzed is supplied to the mass spectrometer 1 from, for example, a liquid chromatography (LC) apparatus such as an LC column (not shown in FIG. 1). One such example of an LC column is the ProSwift (trademark) integrated column manufactured by Thermo Fisher Scientific, which provides high performance liquid chromatography (HPLC) by forcing the sample through a mobile phase under high pressure through a stationary phase of irregularly shaped or spherical particles that form the stationary phase. In an HPLC column, sample molecules elute at different rates depending on the degree of interaction with the stationary phase. The chromatogram may be generated by measuring, over time, the amount of sample molecules eluting from the HPLC column using a detector that is a mass spectrometer.

[0060] Sample molecules eluting from the HPLC column are detected as peaks above the baseline measurement of the chromatograph. If different sample molecules have different elution rates, multiple peaks on the chromatograph may be detected. Preferably, the individual sample peaks are temporally separated from the other peaks on the chromatograph so that different sample molecules do not interfere with each other. In a chromatograph, the presence of a peak on the chromatograph corresponds to the period during which the sample molecule is present in the detector. Thus, the width of the peak on the chromatograph corresponds to the period during which the sample molecule is present in the detector. Preferably, the peak on the chromatograph has a Gaussian-shaped profile or can be assumed to have a Gaussian-shaped profile. Thus, the width of the peak on the chromatograph can be determined based on several standard deviations calculated from the peak. For example, the peak width may be calculated based on four standard deviations of the peak on the chromatograph. Alternatively, the peak width may be calculated based on the width at half the maximum height of the peak. Other methods for determining peak width well known in the art may also be suitable in some cases. Thus, the sample molecules separated by liquid chromatography are then fed into the ion source 11 of the mass spectrometer.

[0061] Here, with reference to FIG. 2, a detailed example of the double analyzer mass spectrometer of FIG. 1 will be described. The double analyzer mass spectrometer 10 of FIG. 2 includes an electrospray ionization source (ESI source) 20 at atmospheric pressure as an ion source. For example, sample molecules received from the HPLC column are ionized by the ESI source 20. Next, the ions generated from the sample enter the vacuum chamber of the mass spectrometer 10 and are directed by the capillary 25 to the RF-only S lens 30. The ions are focused by the S lens 30 into the injection frittapole 40 and inject the ions into the bending frittapole 50 having an axial field. The bending frittapole 50 guides the (charged) ions along a curved path through it, but unwanted neutral molecules such as entrained solvent molecules are not guided along the curved path and are lost.

[0062] The ion gate (TK lens) 60 is located at the distal end of the bent frustoconical pole 50. The ion gate may be an ion lens having static fields that provide good fringe field characteristics and efficiently transmit ions to the quadrupole mass filter 70. In some embodiments, the ion gate may control the passage of ions from the bent frustoconical pole 50 to the quadrupole mass filter 70. The quadrupole mass filter 70 is typically, but not necessarily, segmented and functions as a bandpass filter, allowing the passage of selected mass numbers or a limited mass range while excluding ions of other mass-to-charge ratios (m / z). The mass filter may also be operated in a non-mass-selective mode, i.e., transmitting substantially all m / z ions. For example, the quadrupole mass filter 70 may be controlled by a controller 195 to select the range of mass-to-charge ratios for the passage of precursor ions to be passed, while other ions in the precursor ion stream are filtered (i.e., not passed). Alternatively, the S lens 30 may be operated as the ion gate and the ion gate (TK lens) 60 may be an electric field lens.

[0063] Although a quadrupole mass filter is shown in FIG. 2, those skilled in the art will understand that other types of mass selection devices may also be suitable for selecting precursor ions within the desired mass range. For example, an ion separator as described in U.S. Patent No. A2015287585, an ion trap as described in International Publication No. 2013 / 076307(A), an ion mobility separator as described in U.S. Patent No. 2012 / 256083(A), an ion gate mass selection device as described in International Publication No. 2012 / 175517(A), or a charged particle trap as described in U.S. Patent No. 799223, the contents of which are hereby incorporated by reference in their entirety. Those skilled in the art will understand that other methods of selecting precursor ions according to ion mobility, differential mobility, and / or transverse modulation may also be suitable.

[0064] Isolation of multiple ions of different masses or mass ranges may also be performed using a method well-known as Synchronous Precursor Scanning (SPS) in an ion trap. Further, in some embodiments, two or more ion selection devices or mass selection devices may be provided. For example, a further mass selection device may be provided downstream of the fragmentation chamber 120. In this way, if desired, an MS 3 scan or an MS n scan can be performed (typically, a TOF mass analyzer is used for mass spectrometry).

[0065] Following mass selection, the ions pass through the quadrupole exit lens configuration / split lens configuration 80 and enter the first transfer multipole 90. The quadrupole exit lens configuration / split lens configuration 80 may be used to control the entry of ions into the mass analyzer. The first transfer multipole 90 guides the mass-filtered ions from the quadrupole mass filter 70 into the curved linear ion trap (C-trap) 100. The C-trap (first ion trap) 100 has a longitudinally extending curved electrode to which an RF voltage is supplied and end caps to which a DC voltage is supplied. As a result, a potential well is obtained that extends along the curved longitudinal axis of the C-trap 100. In the first mode of operation, since the DC end cap voltage is set for the C-trap, the ions arriving from the first transfer multipole 90 are trapped in the potential well of the C-trap 100 and cooled there. The injection time (IT) of ions into the C-trap then determines the number of ions (ion population) released from the C-trap into the mass analyzer.

[0066] The cooled ions are trapped in a cloud towards the bottom of the potential well and then are emitted at right angles from the C trap towards the first mass spectrometer 110. As shown in FIG. 2, the first mass spectrometer is an orbitrap type mass spectrometer 110, for example, an Orbitrap (registered trademark) mass spectrometer sold by Thermo Fisher Scientific. The orbitrap type mass spectrometer 110 has an off-centered injection opening, and ions are injected into the orbitrap type mass spectrometer 110 as an agglomerated packet through the off-centered injection opening. Next, the ions are trapped in the orbitrap type mass spectrometer by an ultra-logarithmic electric field and undergo longitudinal back-and-forth motion while orbiting around the inner electrode.

[0067] The axial (z) component of the motion of the ion packet in the orbitrap type mass spectrometer is defined as (more or less) a simple oscillation, and the angular oscillation frequency around the z-axis direction is related to the square root of the mass-to-charge ratio of a given ion species. Therefore, the ions are separated over time according to the mass-to-charge ratio.

[0068] The ions in the orbitrap type mass spectrometer 110 are detected using an image detector (not shown) that generates a "transient" in the time domain containing information about all ion species when passing through the image detector. Next, the transient is subjected to a fast Fourier transform (FFT), resulting in a series of peaks in the frequency domain. A mass spectrum representing the presence / ion intensity for m / z can be generated from these peaks.

[0069] In the above configuration, the sample ions (more specifically, the mass range segment of the sample ions within the target mass range selected by the quadrupole mass filter 70) are analyzed by the orbitrap type mass spectrometer without fragmentation. The resulting mass spectrum is denoted as MS1.

[0070] When it is desired to perform an MS2 scan using an orbital trap type analyzer, the mass spectrometer operates in a second mode where it first needs to fragment the ions. In such a case, the ions are delivered through the fragmentation chamber 120 for cooling and fragmentation. The fragmented ions are then reciprocated back to the C trap 100 where they are pulse extracted into the orbital trap type analyzer 110 for mass analysis.

[0071] In this second operating mode, the ions that enter the C trap 100 through the quadrupole exit lens configuration / split lens configuration 80 and the first transfer multipole 90 continue their path through the C trap and into the fragmentation chamber 120. Thus, the C trap effectively operates as an ion guide in the second operating mode. The fragmentation chamber 120 is a higher energy collisional dissociation (HCD) device in the mass spectrometer 10 of FIG. 2 where a collision gas such as an inert gas like argon, nitrogen or helium is supplied. The precursor ions arriving at the fragmentation chamber 120 collide with the collision gas molecules, as a result of which the precursor ions are fragmented into fragment ions.

[0072] Although the HCD fragmentation chamber 120 is shown in FIG. 2, alternatively, other fragmentation devices using methods such as collision induced dissociation (CID), electron capture dissociation (ECD), electron transfer dissociation (ETD), photodissociation etc. may be used.

[0073] As described above, for MS2 analysis by the orbital trap type analyzer 110, the fragmented ions are returned to the C-trap 100 and reciprocated, and are directed laterally with respect to the C-trap axis toward the orbital trap type analyzer 110. Alternatively, the MS2 analysis may be performed by a second mass analyzer, which may be a time-of-flight mass analyzer 150, as shown in the embodiment of FIG. 2. In such a case, the fragmented ions may be emitted from the fragmentation chamber 120 at the axial end opposite to the C-trap 100. The emitted fragmented ions pass through the second transfer multipole 130. The second transfer multipole 130 guides the fragmented ions from the fragmentation chamber 120 into an extraction trap (second ion trap) 140. The extraction trap 140 is a high-frequency voltage-controlled trap containing a buffer gas. For example, a suitable buffer gas is argon at a pressure in the range of 5×10 -4 mBar to 1×10 -2 mBar. The extraction trap has the ability to quickly switch off the applied RF voltage and apply a DC voltage to extract the trapped ions. A suitable flat plate extraction trap, also referred to as a linear ion trap, is further described in U.S. Patent No. 9,548,195 (B2). Alternatively, the C-trap may also be suitable for use as the second ion trap.

[0074] The extraction trap 140 is provided to form an ion packet of the fragmented ions prior to injection into the second analyzer. The extraction trap 140 accumulates the fragmented ions before injecting them into the time-of-flight mass analyzer 150.

[0075] An extraction trap (ion trap) is shown in the embodiment of FIG. 2, but those skilled in the art will understand that other methods of forming ion packets of fragmented ions are equally suitable for the present invention. For example, the relatively slow movement of ions through a multipole can be used to affect the clustering of ions, which can then be released as a single packet into a ToF mass spectrometer. Alternatively, orthogonal displacement of ions may be used to form packets. Further details of these alternatives are described in U.S. Patent No. 2003 / 0001088 (A1), which describes traveling wave ion bunching methods, the contents of which are incorporated herein by reference.

[0076] In FIG. 2, the time-of-flight mass spectrometer 150 shown is a multi-reflection time-of-flight mass spectrometer (MR-ToF) 150. The MR-ToF 150 is constructed around two opposing ion mirrors 160, 162 that are elongated in the drift direction. The mirrors face each other in a direction orthogonal to the drift direction. The extraction trap 140 injects ions into the first mirror 160, and then the ions oscillate between the two mirrors 160, 162. The angles of ion emission from the extraction trap 140 and the additional deflectors 170, 172 allow control of the energy of the ions in the drift direction, such that as the ions oscillate, they are directed downward along the length of the mirrors 160, 162 to generate a zigzag trajectory. The mirrors 160, 162 themselves are tilted relative to each other to generate a potential gradient that slows the drift velocity of the ions, reflects the ions into the drift dimension, and focuses them onto the detector 180. The tilt of the opposing mirrors typically has the negative side effect of changing the period of the ion oscillation as the ion oscillation moves through the drift dimension. This is corrected by stripe electrodes 190 (functioning as compensation electrodes) that change the length of the opposing mirrors 160, 162 downward to change the flight potential for a portion of the space between the mirrors. The combination of the varying width of the stripe electrodes 190 and the change in the distance between the mirrors 160, 162 allows for maintaining ion reflection and spatial focusing onto the detector 180, as well as good temporal focusing. A suitable MR-ToF analyzer 150 for use in the present invention is further described in U.S. Patent No. 2015 / 028197 (A1), the content of which is hereby incorporated by reference in its entirety.

[0077] The above describes several operation modes. First, it was explained that the MS1 scan can be performed by the first mass spectrometer (Orbitrap mass spectrometer 110). Second, the inventors have determined that depending on whether the fragmentation chamber is controlled to emit ions rearward toward the C-trap 100 or forward toward the second transfer multipole 130, the precursor ions can be fragmented by the first mass spectrometer (Orbitrap mass spectrometer 110) or the second mass spectrometer (time-of-flight mass spectrometer), and the MS2 scan can be performed. In a further operation mode, the second mass spectrometer (time-of-flight mass spectrometer 150) may perform an MS1 scan of the ions. In this operation mode, the ions are axially directed through the C-trap 100 toward the fragmentation chamber, but no fragmentation gas is input, and the ions are induced to the second transfer multipole 130 without fragmentation. Next, the ions can be accumulated in packets within the extraction trap 140 as described above.

[0078] Returning to FIG. 2, when a predetermined number of ions are accumulated in the extraction trap, the ions accumulated in the extraction trap are injected into the MR-ToF analyzer 150 as an ion packet. By ensuring that each packet of ions injected into the MR-ToF 150 has at least a predetermined (minimum) number of ions, the resulting packet of ions reaching the detector will represent the entire mass range of interest in the MS1 spectrum or MS2 spectrum. A single packet of precursor ions or fragmented ions is sufficient, respectively, for the acquisition of the MS1 spectrum or MS2 spectrum. For MS2, this represents an increased sensitivity compared to the conventional acquisition of time-of-flight spectra that are typically acquired and summed for each given mass range segment. Preferably, the minimum total ion current (TIC) within each mass window is accumulated in the extraction trap prior to injection into the time-of-flight mass analyzer. For example, at least N spectra (scans) per second are acquired in the MS2 region by a time-of-flight mass analyzer, where N = 50, or more preferably, 100, or 200, or more.

[0079] Preferably, at least X% of the MS2 scans contain more than Y ions (where X = 30, or 50, or 70, or most preferably 90 or more, and Y = 200, or 500, or 1000, or 2000, or 3000, or 5000 or more). Most preferably, at least 90% of the MS2 scans contain more than 500 ions, or more preferably more than 1000 ions, and ideally more than 5000 ions. This provides an increased dynamic range for the MS2 spectra. The desired number of ions for each MS2 scan may be provided by adjusting the number of ions contained in each packet of fragmented ions. For example, in the embodiment of FIG. 2, the accumulation time of the extraction trap may be adjusted to ensure that a sufficient number of ions are accumulated. Thus, the controller 195 may be configured to determine that a suitable packet of fragmented ions has been formed when either a predetermined number of ions are present in the extraction trap or a predetermined period of time has elapsed. The predetermined period of time may be specified to ensure that the time-of-flight mass spectrometer operates at the desired frequency when the flow of ions into the extraction trap is relatively low.

[0080] The mass spectrometer 10 is under the control of a controller 195 configured to control, for example, the timing of the release of capture components, set suitable potentials on electrodes such as quadrupoles to focus and filter ions, capture mass spectral data from the orbitrap device 110, and capture mass spectral data from the MR-ToF 150, such as by controlling sequences of MS1 and MS2 scans. It will be understood that the controller 195 may comprise a computer operable according to a computer program including instructions to cause the mass spectrometer to perform the steps of the method according to the present invention.

[0081] It should be understood that the particular arrangement of components shown in FIG. 2 is not essential to the method described later. Indeed, other configurations for implementing the method of embodiments of the present invention are also suitable.

[0082] Although an Orbitrap mass spectrometer 110 is shown in FIG. 2, other Fourier transform mass spectrometers may be used instead. For example, a Fourier transform ion cyclotron resonance (FTICR) mass spectrometer may be utilized as the mass spectrometer for MS1 scans. Even when other types of signal processing other than Fourier transform are used to obtain mass spectral information from transient signals, mass spectrometers such as Orbitrap mass spectrometers and ion cyclotron resonance mass spectrometers may also be used in the present invention (see, for example, International Publication No. 2013 / 171313).

[0083] Here, referring to FIG. 3, a representative method in which sample molecules are supplied from a liquid chromatography (LC) column to the above-described representative apparatus (shown in FIG. 2) will be described. Sample ions are supplied from the LC column to acquire data about the sample. The data is acquired over the entire elution period, which is usually controlled by the length of the solvent gradient (switching from pumping an aqueous solvent onto the column to an organic solvent). Mass scans may be collected over repeated cycles. It is preferable that each scan or scan cycle for a target mass range or list of precursors is performed over a duration or time scale corresponding to the elution of a chromatographic peak or less than that.

[0084] As shown in FIG. 3, an Orbitrap mass spectrometer (denoted as "Orbitrap") is utilized to perform a plurality of MS1 scans over a target mass range. For example, as shown in FIG. 3, the target mass range to be analyzed (or selected ion monitoring, SIM) is 400 to 1000 m / z. Within the target mass range, a plurality of MS1 scans are performed using mass sub-ranges of the precursor ions in the target mass range. Alternatively, a single MS1 scan may be performed using precursor ions from the entire target mass range (i.e., in this example, 400 to 1000 amu).

[0085] To perform a single MS1 scan, sample molecules from the LC column are ionized using the ESI source 20. Thereafter, the sample ions enter the vacuum chamber of the mass spectrometer 10. The sample ions are directed through the capillary 25, the RF-only lens 30, the injection frustum 40, the bending frustum 50, and as described above, towards the quadrupole mass filter 70. The quadrupole mass filter 70 is controlled by the controller 195 and filters the sample ions according to the selected precursor mass sub-range of interest. For example, as shown in FIG. 3, the MS1 scan is performed over a target mass range of 400 m / z to 1000 m / z within precursor mass sub-ranges of 400 - 500 m / z, 500 - 600 m / z to 900 - 1000 m / z. Next, the ions pass through the quadrupole exit lens configuration / split lens configuration 80, through the transfer multipole 90, and enter the C-trap 100 where they are accumulated. From the C-trap 100, (precursor) sample ions of a mass range segment may be injected into the orbitrap mass analyzer 110. When the ions are stabilized inside the orbitrap mass analyzer, an MS1 scan is performed by using an image current detector to detect the ions present in the orbitrap mass analyzer 110. The detection of ions in the orbitrap mass analyzer is configured to be performed with a relatively high resolution for the MS1 scan (compared to the resolution of the MS2 scan). For example, a resolution (R) of at least 50,000, or preferably at least 100,000, may be used for each MS1 scan (see resolution R = 120,000 in FIG. 3).

[0086] By using a Fourier transform mass analyzer (e.g., an orbitrap mass analyzer), the MS1 scan is performed with a high mass accuracy. Preferably, the MS1 scan is performed with a mass accuracy of less than 5, or more preferably, less than 3 parts per million (ppm).

[0087] As shown in FIG. 3, a plurality of MS2 scans are performed in parallel with the MS1 scan. In the method of FIG. 3, the MS2 scan is performed using a time-of-flight mass spectrometer MR-ToF. In other embodiments, different mass spectrometers may be used.

[0088] To perform a single MS2 scan of a mass range segment, sample molecules from the LC column are ionized and injected into the mass spectrometer in the same manner as in the MS1 scan. Sample ions for the MS2 scan proceed through the capillary 25, RF-only lens 30, entrance frustum 40, and bent frustum 50, and into the quadrupole mass filter 70, just like the sample ions for the MS1 scan. When the sample ions for the MS2 scan reach the quadrupole mass filter 70, the quadrupole mass filter 70 is controlled by the controller 195 to filter the sample ions according to the relatively narrow mass range segment being scanned. Each precursor mass range segment may have a mass range of, for example, 5 amu or less (as shown in FIG. 3), or preferably 3 amu or less, or more preferably 2 amu or less. The (filtered mass range segment) precursor ions pass from the quadrupole mass filter 70 to the C-trap 100 as described above for the MS1 scan. Next, the controller 195 controls the C-trap to allow the precursor ions to pass axially towards the fragmentation chamber 120. In the HCD fragmentation chamber 120, the precursor ions collide with collision gas molecules, and as a result, the precursor ions are fragmented into fragment ions. The fragmented ions for the mass range segment are then released from the fragmentation chamber at the opposite axial end with respect to the C-trap 100. The released fragmented ions pass through the second transfer multipole 130. The second transfer multipole 130 guides the fragmented ions from the fragmentation chamber 120 into the extraction trap (second ion trap) 140, where they are accumulated. The fragmented ions may be accumulated in the extraction trap 140 for a predetermined time. The fragmented ions are then injected from the extraction trap into the MR-ToF. The pre-accumulation of fragmented ions in the extraction trap enables the fragmented ions to be injected into the MR-ToF as a packet. The ion packet moves along the flight path of the MR-ToF and undergoes multiple reflections before being detected by the detector.The various arrival times of the fragmented ions within the packet enable the generation of an MS2 mass spectrum for the packet of fragmented ions. The length of the MR-ToF flight path, combined with the time resolution of the detector, enables the MR-ToF to perform MS2 scans with a resolution exceeding 40000 (see R = 50000 in Figure 3).

[0089] One advantage of the packet-based approach to analysis is that when the accumulated ions are released from the extraction trap 140, ions for the next mass range segment can begin to fill the extraction trap. Thus, ions from one mass range segment can be moved through the MR-ToF while ions for the next mass range segment are being accumulated. Thus, by using packet-based injection of fragmented ions from the extraction trap into the MR-ToF, a greater number of ions can be achieved within the chromatographic peak. This holds true whether an MS1 scan or an MS2 scan is being acquired, or when switching from an MS1 scan to an MS2 scan or vice versa.

[0090] The controller 195 controls the mass spectrometer 10 to perform a plurality of MS1 scans of mass sub-ranges for a target mass range and, in parallel, perform a plurality of MS2 scans of mass range segments across the target mass range. To obtain a more accurate sample of the chromatographic peak, the controller 195 may repeat the scan cycle a plurality of times over the duration of the chromatographic peak. For example, a single cycle may take about 1.5 seconds to perform. Thus, the cycle may be performed at least 7 times, or more preferably at least 9 times, over the duration of the chromatographic peak. This enables MS1 spectral data and / or MS2 spectral data to be used for quantification of the eluted sample at the chromatographic peak. In some operating modes, a single operation of the cycle may be sufficient.

[0091] Figure 3 shows the MS1 acquisition and MS2 acquisition for single polarity. The lower part of the figure shows how ions from the ion source are distributed to two analyzers for MS1 and MS2 acquisition. It can be seen that across the ions from the ion source, the ions from the ion source are directed to the MR-ToF for MS2 acquisition.

[0092] If it is desired to collect MS1 scans and / or MS2 scans across both polarities, as discussed, the HRAM analyzer has a long polarity switching time that results in a dead time where the analyzer(s) cannot perform reliable scans. This is caused by the time it takes for the voltage supply to the mass analyzer to stabilize after polarity switching. There is also some dead time while the ion source and ion induction region are switching polarities, but this is much shorter than the dead time of the analyzer.

[0093] Embodiments of the present invention relate to maximizing the use of ions such as ions eluting from a sample from a liquid chromatography (LC) column and minimizing the temporal deviation in the acquisition of mass scans so as to be able to analyze all target ion species or fragmented ion species. FIGS. 4-8 show a method of performing mass spectrometry using a dual analyzer mass spectrometer according to an embodiment of the present invention. In FIGS. 4-8, the positive polarity operation of the ion source / ion induction and the mass analyzer is shown by a thick black line, and the negative polarity operation of the ion source / ion induction and the mass analyzer is shown by a thin double gray line.

[0094] Embodiments of the present invention include covering the dead time of polarity switching of one analyzer by deflecting an ion beam to a second analyzer. In this way, the ion beam is always utilized efficiently, and only the short polarity switching times of the ion source and the ion induction region create a dead period. For further explanation, when considering the apparatus of FIG. 1 or FIG. 2, the apparatus can be divided into sections, and for example, for the ion source, the ion processing region, the first mass analyzer, and the second mass analyzer, the polarities of each section can be configured independently. In the mass spectrometer of FIG. 2, the ion source corresponds to the ESI source 20, and the ion processing region corresponds to the capillary 25, the RF-only lens 30, the injection frustum 40, the bending frustum 50, the ion gate 60, the quadrupole mass filter 70, the exit lens configuration / split lens configuration 80, the transfer multipole 90, and the C-trap 100. The polarities of the components within the ion processing region and the ion source are preferably switched together. Depending on the scan being performed, the fragmentation chamber 120, the second transfer multipole 130, and the extraction trap 140 may also have their polarities switched by the ion processing region. Further division or sub-division of the switching of the ion source and the ion processing region is possible depending on the implementation and scan being performed. In the embodiment of FIG. 2, the first mass analyzer is an orbitrap-type analyzer 110, and switching the polarity of the orbitrap-type analyzer 110 may include switching the orbitrap-type analyzer and the injection optics (shown between reference numerals 100 and 110 in FIG. 2). Also, in the embodiment of FIG. 2, the second analyzer is an MR-ToF analyzer 150, and switching the polarity of the MR-ToF analyzer may include switching the polarities of items 160-180, and may further include switching the second transfer multipole 130 and the extraction trap (second ion trap) 140. Many of the components within the ion source and the ion induction / processing region switch relatively quickly, but the DC voltage extracted from the C-trap 100 to the orbitrap analyzer may be slower, but is only required for orbitrap analyzer scans.

[0095] The ion source and the ion processing region generally operate with the same polarity. To avoid loss of ion usage while one mass analyzer is switching, the ion beam or ions are switched to the other analyzer. Since different types of analyzers have different switching times, the optimal method for a particular pair of analyzers depends on their type and switching times.

[0096] In FIGS. 2 and 3, the inventors considered a dual mass analyzer instrument where the first analyzer is an Orbitrap (trademark) analyzer and the second analyzer is a multi-reflection time-of-flight mass analyzer (MR-ToF).

[0097] As a first embodiment, consider the switching of an instrument where the first analyzer is an Orbitrap analyzer and the second analyzer is an ion trap. Such an instrument is equivalent to Orbitrap (trademark) Exploris (trademark). The Orbitrap analyzer may have a polarity switching time of about 500 ms. This is a relatively long switching time, but it is compatible with the chromatographic peak elution time scale. The Orbitrap analyzer requires a significant amount of ion processing time to generate high-quality full mass scans. The ion trap is highly sensitive and switches polarity more rapidly, in about 20 ms. For MS1 spectra, a mass analyzer with high resolution and mass accuracy is required, while for MS2 spectra, high sensitivity and speed are required. FIG. 4 shows the timeline of a dual mass analyzer instrument using an Orbitrap analyzer for MS1 scans and an ion trap for MS2 scans.

[0098] As shown in Figure 4, the two polarities are covered by having a first phase in which the ion source, the ion guiding / processing region, the first mass analyzer (orbitrap analyzer), and the second mass analyzer (ion trap) all operate in a first polarity that is positive. The term "operating in the first polarity" means that each of the ion source, the ion guiding / processing region, the first mass analyzer, and the second mass analyzer is configured to operate for the analysis of ions in the first polarity. For example, in the case of the ion source, this can mean that the nebulizer is set to a high positive voltage (+4KV) to generate positive ions, and the downstream electrode has a wide negative DC voltage progression to facilitate the movement of positive ions through them. The applied RF potential that provides radial focusing is essentially polarity independent. Following the first step of operating in the first positive polarity, in the second step, the ion source, the ion guiding / processing region, the first mass analyzer (orbitrap analyzer), and the second mass analyzer (ion trap) all operate in a second negative polarity. When the second phase is complete, the polarities of the ion source, the ion guiding / processing region, the first mass analyzer (orbitrap analyzer), and the second mass analyzer (ion trap) may be switched back to the first polarity, i.e., positive, to operate. Alternatively, the first polarity may be negative and the second polarity may be positive.

[0099] Looking in more detail at the timing for switching polarities in FIG. 4, the dead time (DT1) for switching the polarity of the orbitrap analyzer is used by the ion trap for MS2 acquisition, while the dead time for switching the ion trap is used by the orbitrap mass analyzer for MS1 acquisition. The orbitrap analyzer loaded with ions from the ion source takes a relatively long time to generate the MS1 spectrum, during which time, after the ion trap has switched polarity, acquisition of the MS2 spectrum can be initiated. Multiple MS2 spectra may be collected while the MS1 spectrum is being generated. After the orbitrap analyzer has completed MS1 acquisition, the polarity of the orbitrap analyzer may be switched so that the ion source and the ion guiding / processing region are also prepared as if their polarities have been switched. The ion trap continues in the positive mode of collecting the MS2 spectrum during this dead time of the orbitrap analyzer switch. In this way, the ions from the ion source can be utilized to the maximum extent. The collection of ions and the generation of the MS2 spectrum continue using the ion source and the ion guiding / processing region with the ion trap operating in the first polarity, i.e., the positive polarity, until the final MS2 spectrum is initiated. At this point, after the ions have been collected for the final MS2 spectrum in the first polarity, the ion source and the ion guiding / processing region are switched to the second polarity. The ion trap continues to process the final MS2 spectrum in the first polarity during the relatively short dead time (DT2) for switching the polarity of the ion guiding and processing region. After the ion source and the ion guiding / processing region have switched polarity and the orbitrap analyzer has also switched polarity, acquisition of the MS1 spectrum in the second polarity may be initiated. The ion trap may complete the final MS2 acquisition while the orbitrap analyzer is starting its MS1 acquisition in the second polarity which is negative. Next, the process is repeated in the second polarity using an MS2 scan starting from the ion trap after ions for the MS1 scan have been trapped.The switching polarity of the ion trap also has a dead time (DT3) during which the ion trap is not available for use. Advantageously, this dead time is timed to coincide with the acquisition of ions by the orbitrap analyzer for MS1 scanning so that the use of the ion / ion beam is maintained.

[0100] Accordingly, FIG. 4 shows the instrument cycle in which the second analyzer (ion trap) is performing MS2 acquisition when the first analyzer (orbitrap analyzer) is either acquiring MS1 or switching polarity, and the first analyzer (orbitrap analyzer) is loading ions for MS1 acquisition when the second analyzer (ion trap) is switching polarity. The ion source may also switch polarity while the ion trap is performing its last analysis. As can be seen from the figure, the dead time (DT) repeats when the ion source and the ion processing region / ion induction, the first analyzer, and the second analyzer switch polarity a second time, i.e., return to the first polarity. There is also a dead time at the start of the timeline of the figure for the second analyzer when the second analyzer switches to the first polarity while the orbitrap analyzer is loading ions. If the start of the timeline in FIG. 4 represents the first acquisition from the sample, the second analyzer may already be set to the correct polarity from the start and may not need to switch initially.

[0101] Figure 4 shows a method applied to an orbitrap type analyzer as the first mass analyzer and to an ion trap as the second mass analyzer, although other types of analyzers may be used. Further, Figure 4 shows an orbitrap type analyzer where the ion source performs the first acquisition after switching the polarity, although the sequence may be reversed so that the ion trap performs the first acquisition. In such a case, the orbitrap type analyzer switching time may occupy the initial portion of the time at the first polarity. Compared to the method of Figure 4, this method is not very suitable for data-dependent analysis (DDA) because the MS1 data is not collected and processed before the MS2 scan, and thus cannot be used to determine the m / z mass range in which the MS2 spectrum should be collected. However, since the cycle should be repeated many times for each experiment, only a part of the first cycle may be lost, and subsequent cycles provide data similar to the method of Figure 4.

[0102] A second embodiment similar to the first embodiment is shown in Figure 5. Here, the second analyzer is a time-of-flight analyzer (ToF) such as the MR-ToF of Figures 2 and 3. The timing and operation of the ion source and the ion induction / ion processing region are substantially the same as those described for Figure 4. However, the ToF analyzer may have a longer polarity switching time. Thus, as shown in Figure 5, the increased polarity switching time of the ToF analyzer compared to the ion trap of Figure 4 may result in the MS2 scan starting slightly later and fewer MS2 scans being performed. Nevertheless, as in Figure 4, Figure 5 shows that the second analyzer (ToF) performs MS2 acquisition when the first analyzer (orbitrap type analyzer) is either acquiring MS1 or switching polarity, and when the second analyzer (ToF) is switching polarity, the first analyzer (orbitrap type analyzer) is loading ions for MS1 acquisition and optionally starting MS1 acquisition. The ToF may switch polarity while the ToF is performing its last analysis.

[0103] The ToF analyzer as the second analyzer has a longer switching time and reduced operation time compared to the ion trap of FIG. 4. However, for example, if the ToF analyzer is MR-ToF and highly sensitive, and thus suitable for MS2 scanning, it is still preferable to use an orbitrap-type analyzer as the first analyzer for performing MS1 scanning. Furthermore, high mass accuracy may not be as important for ToF MS2 scanning, and the long electron stabilization time of the electronics may be slightly shortened.

[0104] Figure 6 shows a third embodiment. This embodiment is the same as Figure 5 in that the mass spectrometer includes an orbitrap type analyzer as the first analyzer and a ToF analyzer as the second analyzer. Different from Figure 5, in Figure 6, the polarity of the ToF analyzer is fixed and it is only used for MS2 scans in one polarity. In Figure 6, the ToF analyzer is only used for positive polarity (alternatively, this may be only negative polarity). The orbitrap type analyzer performs MS1 scans of both polarities and negative polarity MS2 scans. The method in Figure 6 again uses the idea of reducing the impact of the switching dead time of one analyzer by directing ions from the ion source to the second analyzer to perform scans. Here, the method in Figure 6 will be described in more detail. Different from Figures 4 and 5 where the ion source and the ion guiding / ion processing region require two polarity switches to complete all cycles of positive and negative MS1 and MS2 scans, Figure 6 requires the polarity to be switched four times. The cycle starts in the same way as in Figure 5, where the orbitrap type analyzer collects ions for a positive polarity MS1 scan, and then the ToF analyzer generates a positive polarity MS2 scan. The time when the orbitrap type analyzer processes ions to obtain an MS1 scan is filled by the operation of the ToF that collects the MS2 scan, thereby maximizing the use of ions / ion beams. After the orbitrap type analyzer completes the MS1 scan, it starts a polarity switch (DT1’), while the ToF analyzer continues to collect the MS2 spectrum. The next step is also the same as the step in Figure 5 in that after ions are collected for the last of a group of MS2 scans, the polarity of the ion source and the ion guiding / ion processing region is switched. When the ion source / ion processing region switches polarity, the orbitrap type analyzer is also switched, and the acquisition of a negative MS1 scan by the orbitrap type analyzer starts. The next step is different from Figure 5. After the orbitrap type analyzer collects ions to generate a negative MS1 scan, the ion source and the ion guiding / ion processing region are switched back to the first polarity, i.e., positive polarity, and the ToF can collect more positive MS2 scans.Here, as soon as the ion source and the ion guiding / ion processing region are switched, since the ToF is already in the required polarity, MS2 acquisition can be started. After the negative MS1 scan is completed, the ion source and the ion guiding / ion processing region are switched to negative, enabling the generation of a negative MS2 scan by the orbitrap type analyzer. The orbitrap type analyzer is already in the negative polarity just after completing the negative MS1 scan, and thus can start negative MS2 acquisition as soon as the ion source and the ion guiding / ion processing region are switched. Also, by performing the negative MS1 and negative MS2 scans successively, the number of times the orbitrap type analyzer is switched in this method is minimized. After the ions for the last negative MS2 scan by the orbitrap type analyzer are acquired, the ion source and the ion guiding / ion processing region are switched in polarity, and the acquisition of a positive polarity MS2 scan can be started. While these scans are being generated, the orbitrap type analyzer is switched in polarity to return to the first polarity, i.e., the positive polarity.

[0105] In the method of FIG. 6, although additional switching is required, the significant dead time that may occur due to the long switching time of the ToF analyzer is avoided. While the orbitrap type analyzer generates an anion MS2 spectrum, there is a deviation in the ToF operation, which means that there is a period during which positive MS2 ions are not analyzed and the elution of species from the LC column may be overlooked. However, when the MS2 acquisition time is longer, it may be possible to quickly switch the source polarity and acquire additional positive MS2 ToF spectra interleaved with the negative orbitrap type analyzer MS2 spectra. Further, in the scheme shown in FIG. 6, more results are obtained for the positive polarity side of the measurement. If equivalent results are desired for the negative polarity side, the time allowance for each scan can be adjusted to equalize the weighting.

[0106] The fourth embodiment is shown in FIG. 7. Similar to FIGS. 5 and 6, this embodiment also uses an orbitrap type analyzer as the first analyzer and a ToF analyzer as the second analyzer. In FIG. 7, the switching amount is reduced. This is achieved by using a ToF or MR-ToF analyzer of sufficient quality for MS1 acquisition. For example, a ToF or MR-ToF analyzer having at least >30K resolution and <10ppm mass accuracy, which can be provided by the latest qToF, would be sufficient. However, >50K resolution and <5ppm mass accuracy are preferred. In such cases, there is no need to switch the polarity of either analyzer. Only the ion source and the ion induction / ion processing region need to be switched. In the scheme shown in FIG. 7, the ion source and the ion induction / ion processing region are initially shown in the negative polarity, and the orbitrap type analyzer is generating a negative MS2 spectrum. During this time, the ToF analyzer is not used and an idle time is experienced. After sufficient negative MS2 spectra have been collected by the orbitrap type analyzer, ions are collected so that the orbitrap type analyzer generates an MS1 scan. After the ions are collected, the ion source and the ion induction / ion processing region are switched to the second polarity, i.e., the positive polarity. Next, positive MS1 acquisition by the ToF analyzer can be started, and then positive MS2 scans by the ToF analyzer follow. The positive MS1 and positive MS2 scans by the ToF analyzer are approximately simultaneous with the orbitrap type analyzer that generates a negative MS1 scan. After the orbitrap type analyzer completes the negative MS1 scan and the ToF analyzer completes the positive MS2 scan, the polarity of the ion source and the ion induction / ion processing region is switched back to the first polarity, i.e., the negative polarity. Those skilled in the art will understand that the corresponding method may be implemented with the first and second polarities reversed. Also, the timing of the MS1 and MS2 scans may be reversed for, e.g., the positive scans performed by the ToF analyzer. As shown in FIG. 7, when MS1 acquisition is performed by the ToF analyzer, this may be done in a plurality of MS1 scans each directed to a mass sub-range of the target mass range (as described above with respect to FIG. 3).Alternatively, a single MS1 scan may be performed over the entire target mass range.

[0107] (As described in connection with FIGS. 4 and 5) There is no polarity switching of one analyzer for another analyzer to cover, but the scheme of FIG. 7 uses the long orbitrap type analyzer MS1 acquisition time to acquire the ToF MS1 and MS2 spectra. Thus, the potential dead time is minimized again by operating a combination of hybrid analyzers. Again, the percentage of time the ion source spends in negative or positive mode may be varied, for example, if more time is required to collect the orbitrap type analyzer MS2 spectrum than an equivalent ToF spectrum.

[0108] A fifth embodiment is shown in FIG. 8. This scheme uses a dual orbitrap type analyzer instrument, i.e., has an orbitrap type analyzer as the first analyzer and a second orbitrap type analyzer as the second analyzer. Here, each orbitrap type analyzer is dedicated to one polarity. Thus, as shown in FIG. 8, the first analyzer is set to negative polarity and the second analyzer is set to positive polarity. The ion source and the ion guiding / ion processing region cycle from one polarity to the other, directing ions to the respective analyzers between each polarity.

[0109] In FIG. 8, the ion source and the ion guiding / ion processing region start in positive polarity and the second orbitrap type analyzer is generating a positive MS2 scan. Meanwhile, the first orbitrap type analyzer receives ions from the ion source and the ion processing region while it is in negative polarity before switching to positive polarity and performs a negative MS1 scan. After the second orbitrap type analyzer completes the positive MS2 scan, the second orbitrap type analyzer receives ions to start a positive MS1 scan. After ions are provided for the positive MS1 scan, the ion source and the ion processing region are switched to negative polarity and the first orbitrap type analyzer generates a negative MS2 scan. The negative MS2 scan is performed by the first orbitrap type analyzer and the positive MS1 scan is performed by the second orbitrap type analyzer.

[0110] In summary, the scheme of FIG. 8 induces ions in one analyzer (at the first polarity) to obtain a series of MS2 spectra, while the other analyzer processes MS1 acquisition in parallel (at the second polarity). This scheme has a minimal dead time and equal time provided for both polarities due to the fact that the orbitrap type analyzer does not need to switch polarities. It may be desirable to provide more time for the MS2 spectrum than for the MS1 spectrum. This can be achieved by extending the time for one polarity when more MS2 scans of one polarity are desired, and the MS1 transient length at the other polarity can be extended to eliminate the dead time.

[0111] The present invention also provides a computer-implemented method for controlling a dual-analyzer mass spectrometer for mixed polarity operation, as shown in FIG. 9. This method may be implemented by a controller such as controller 195 of FIG. 2, or may be implemented by a computer in combination with controller 195.

[0112] The method may cause the computer and / or controller to automatically perform scans, select optimized timings for performing bipolar MS1 and MS2 scans, or provide input requests, or receive input from the user in a user interface. The computer and / or controller may store the time (T ps,A1 , Tps,A2 ) required to switch the polarity for the first and second analyzers, and similarly the time (T ps,IG ) required to switch the polarity for the ion source and ion induction region of the mass spectrometer. These times may be stored in a memory or database 210.

[0113] In one embodiment, the computer / controller may divide the control of the polarity of the ion induction region for simultaneous multiple ion packet processing. Here, the computer / controller selects the polarity of most of the components in the induction region based on the ions / packets that pass through them immediately. Most of the components in the processing region switch their polarities individually very quickly, so there is no extra inoperative time. The computer / controller knows the polarity that each component must have at each time when the ions arrive (e.g., based on the ion migration time) and is programmed to process several ion packets simultaneously. For example, for the simultaneous processing of cations and anions as described with respect to FIGS. 4 to 8, the controller may control the extraction trap 140 to hold the cations, while the quadrupole 70 is controlled to accept the anions.

[0114] As described above with respect to FIGS. 4-8, there are several ways to optimally acquire both polarities of MS1 and MS2 scans. The optimal method(s) for a given mass spectrometer depends on the type of mass analyzer used and their respective switching times. For some types of analyzers, such as an MR-ToF analyzer, it may be preferable to fix the analyzer to one polarity. This may be determined automatically by a computer or controller, as indicated by 220, or may be set by the user. In some methods, such as those shown in FIGS. 7 and 8, it may be preferable to fix the polarity of both analyzers. The user may also desire to set whether the acquisition mode is a data-independent acquisition (DIA) mode in which the mass spectrometer cycles through a preset list of MS1 and MS2 acquisitions in the specified order, or the user may set the acquisition mode to a data-dependent acquisition (DDA) mode in which the MS1 scan is used to generate a precursor list and that list is used to perform a series of MS2 scans of fragment ions based on the precursor list. The DDA method is more focused, but the DIA method may be better when little is known about the species being analyzed. Preferably, the choice between the DIA mode and the DDA mode is made by the user, as shown at 230 in FIG. 9(b).

[0115] In operation 240, the computer and / or controller selects from the available methods based on the information at 210, 220, and 230. For example, depending on the type of mass spectrometer and the user's selection of DIA or DDA, some methods may not be available. Once it is determined which methods are available, the computer may determine the timing and duration for performing MS1 scans and MS2 scans by each analyzer, which may include calculating the amount of ion beam dead time at 250 during which the ion beam is not directed at the analyzer, and checking for overlap of analyzer operation at 260 so that the analyzers are used maximally. Next, the computer and / or controller configures the mass spectrometer to perform MS1 and MS2 scans in bipolarity in operation 270 based on inputs 210, 220, 230, and the available methods. Optionally, the computer and / or controller may generate a warning at 280 if the ion beam is not used for a predetermined amount of time or a predetermined percentage of time.

[0116] When determining the timing of MS1 and MS2 scans, the controller and / or computer may take into account the length of time for performing an MS1 scan by each analyzer and the length of time for performing an MS2 scan by each analyzer. For example, the controller and / or computer may determine that while a single MS1 scan is being performed by one of the analyzers, the other analyzer(s) may have time to perform X number of MS2 scans. The computer and / or controller may also identify when one or more scans are completed, such as after an MS1 scan or after a chain of MS2 scans, etc., and polarity switching may be required such that the polarity switching time(s) of one or both of the analyzers that switch polarity and / or the polarity switching time of the ion source and ion induction region can be taken into account. Preferably, the timing for performing the scans may include, as necessary, the ion source and ion induction region and the ion transfer time from any ion trap.

[0117] Specifically different methods rely on relatively optimized timings of different devices under computer control. For example, if the selected method is based on the method of FIG. 6, the control has to provide a sufficient number of ToF MS2 spectra to fill the dead spots of the orbitrap analyzer polarity switching. The computer-implemented method may calculate the time required for various operations including scanning and polarity switching. For example, in FIG. 6, the MR-ToF cycle takes about 5 ms, but the orbitrap analyzer MS1 scan may require 256 ms for a suitable 120K resolution. As described above, in the data-independent method that generally circulates a preset acquisition list in a preset order, it may be preferable for the user interface to provide the user with the visualization or calculation of the duplication of the analyzer operation and return a warning against wasteful beam time.

[0118] To determine a data-dependent method in which the experimental cycle creates a precursor list from an MS1 scan and then implements that list through a series of N MS2 scans, the method should preferably define that the time allowance for the MS2 scan does not exceed the time for the MS1 scan + polarity switching. It may also be necessary to take into account the priority between positive MS1 and negative MS2. For example, the method of FIG. 6 has negative ion MS2 implemented at a low speed by an orbitrap analyzer, so only 10 negative MS2 scans per cycle may be allowed, but potentially 100 positive ToF MS2 scans per cycle may be allowed.

[0119] The computer-implemented method preferably automatically optimizes this method. For example, when there are fewer anions, the balance of instrument time may be tilted towards a larger positive mode of operation, and vice versa. This preference for one polarity may also be additional input provided by the user before the computer-implemented method determines the preferred analysis method. Algorithm optimization may be necessary for more complex methods, such as special on-the-fly adjustment, multiple MS1 scan ranges, different fragmentation methods with unique processing times, and methods involving variations in the orbitrap analyzer MS1 scan length based on the number (or prediction) of precursors. The computer-implemented method may generate various analysis methods and present them to the user on a user interface, allowing the user to select the preferred analysis method and optionally modify the preferred method.

[0120] The above has described how the time for polarity switching of the mass spectrometer delays the start of the mass spectrometer from the resumption of measurement. This is mainly caused by waiting for the HV adjustment power supply to become sufficiently stable. FIG. 10 schematically shows the voltage fluctuations after the HV voltage source switches polarity. The deviation decreases as time passes. Immediately after switching, the voltage fluctuations are large and ions m / z cannot be measured, but there is an intermediate time when ions m / z can be measured. Next, when the fluctuations are reduced to a low level, ions m / z may be measured to the accuracy required. These different measurement regimes are shown in FIG. 10. It has been determined that it is possible to measure the measurement mass shift associated with the time caused by the fluctuating HV during the intermediate time and, to some extent, immediately after switching. By measuring this variation, a suitable correction may be applied to the ion m / z measurement value to remove the inaccuracies caused by the variation and effectively extend the accurate m / z measurement range closer to the switching time. Similarly, methods of incorporating internal calibration substance correction around the polarity switch, or methods that use only low-accuracy requirement acquisitions such as MS2 scans, also allow for a reduction in the dead time.

[0121] The above described an embodiment of a dual analyzer mass spectrometer with reference to FIG. 2. In an alternative embodiment, the mass spectrometer may be provided with a branched path configuration, for example, as shown in the embodiment of FIG. 11. In the embodiment of FIG. 11, the ion source 400 is coupled to an ion processing region such as a mass selection device 410. Such an arrangement may be provided, for example, by their respective couplings to the ESI ion source 20 and the quadrupole mass filter 70 as shown in the embodiment of FIG. 2.

[0122] As shown in FIG. 11, the output of the mass selection device 410 is coupled to a branched ion path 420. The branched ion path directs the ions output from the mass selection device along one of two paths. The first path 422 guides the ions to a C-trap 430 where the ions are collected for analysis by a first mass spectrometer, for example, an orbitrap-type mass spectrometer 440 within the MS1 domain. The second path 424 directs the ions to a fragmentation chamber 450 for ion fragmentation and subsequent mass spectrometry in the MS2 domain by a second mass spectrometer 470. The branched ion path may use an RF voltage to direct the ions along either the first path 422 or the second path 424. The branched ion path may be a branched RF multipole. A branched ion path suitable for use in the embodiment of FIG. 11 is further described in U.S. Patent No. 7,420,161.

[0123] In the embodiment of FIG. 11, the branched ion path may be used to direct ions to the C-trap 430 for MS1 analysis or to the fragmentation chamber 450 for MS2 analysis. The fragmented ions released from the fragmentation chamber 450 may be accumulated in an ion extraction trap 460 before being injected into the MR-ToF analyzer 470 as a packet. Thus, the arrangement of the fragmentation chamber 450, the ion trap 460, and the MR-ToF 470 may be provided by an arrangement similar to that described in FIG. 2.

[0124] Thus, according to the embodiment of FIG. 11, ions may be directed for MS2 analysis without the need for the C-trap 430 feeding the MS1 orbital trap mass spectrometer 440 to be empty. Such a configuration may allow for increased parallelization of MS1 and MS2 scans. Thus, a greater proportion of the chromatographic peak dwell time may be available for performing MS2 scans. Further, in this configuration, some loading or filling may be accumulated in the C-trap 430 prior to analysis in the orbital trap mass spectrometer 440. In such an embodiment, the loading (filling) of the C-trap 430 may be divided into several small fillings while the orbital trap mass spectrometer 440 is scanning, thereby obtaining a more representative population of ions over the entire peak.

[0125] A further alternative embodiment of an embodiment of a dual analyzer mass spectrometer is shown in FIG. 12. FIG. 12 depicts a schematic diagram of a tandem mass spectrometer 500 including an orbital trap mass spectrometer 510 and a time-of-flight mass spectrometer 520 in a branched path configuration.

[0126] The embodiment of FIG. 12 includes an ion source 530 and an ion guide 540 that supply precursor ions to a mass selector 550. Such an arrangement may be provided, for example, by the respective couplings of an ESI ion source 20 and a quadrupole mass filter 70 as shown in the embodiment of FIG. 2.

[0127] As shown in FIG. 12, a branched ion path 560 directs ions from the mass selector 550 to a C-trap 570 and / or an extraction trap 580. The C-trap 570 supplies ions to the orbital trap mass spectrometer 510 for MS1 scans, and the extraction trap 580 supplies ions to the time-of-flight mass spectrometer 520. For example, a configuration similar to the embodiment disclosed in FIG. 11 may be provided.

[0128] FIG. 12 further includes double linear traps 600, 610. The double linear traps are connected downstream of the C-trap 570 between the C-trap 570 and the extraction trap 580 of the time-of-flight mass spectrometer. The double linear traps may be connected to the C-trap 570 and the extraction trap 580 by ion guides 620, 630. The ion guide 630 may be a branched ion path that merges with the ion path from the mass selector 550 to connect to the extraction trap 580.

[0129] The double linear traps 600, 610 may be provided for ion fragmentation and / or mass separation. For example, the first ion trap 600 may be provided as a high energy collision dissociation chamber. The second ion trap 610 downstream of the first ion trap may be provided as a low collision dissociation chamber. By including the second dissociation chamber, the fragmented ions may be readily fragmented again in the second chamber for MS3 analysis. Ions may be repeatedly isolated and fragmented for MS n analysis. The double linear traps may also enable fragmentation by collision induced dissociation (CID), electron capture dissociation (ECD), electron transfer dissociation (ETD), ultraviolet photo dissociation (UVPD), etc. Further details of a suitable double ion trap can be found in U.S. Patent No. 8,198,580(B), the content of which is incorporated herein by reference in its entirety.

[0130] Advantageously, by providing a direct branch path from the mass selector 550 to the extraction trap 580, ions may be transferred more efficiently from the mass selector 550 to the extraction trap.

[0131] Those skilled in the art will readily understand that various modifications and changes can be made to the above-described methods and apparatuses. Modifications may be made without departing from the scope of the appended claims. For example, the order of the MS1 scan and the MS1 scan, and whether it is negative polarity or positive polarity, may be changed. Steps of methods from different embodiments may be combined. Alternative double analyzer mass spectrometers and alternative mass analyzers may be used.

Claims

1. A method of operating a dual mass spectrometer to obtain MS1 scans and MS2 scans of cations and anions from a sample, comprising: Ionizing the sample in the ion source and ion processing region of the mass spectrometer to generate a plurality of ions, wherein the ion source and the ion processing region operate with a first polarity, generating the plurality of ions; Directing a first ion packet of the plurality of ions to a first mass spectrometer, and performing, by the first mass spectrometer, a first scan sequence including at least one MS1 scan or at least one MS2 scan of the ions in the first ion packet with the first polarity, and after performing the at least one MS1 scan or the at least one MS2 scan, switching the polarity of the first mass spectrometer to a second polarity; Directing one or more second ion packets of the plurality of ions to a second mass spectrometer, and performing, by the second mass spectrometer, a second scan sequence including at least one MS1 scan or at least one MS2 scan of the ions in the one or more second ion packets with the first polarity, wherein at least a part of the at least one MS1 scan or at least a part of the at least one MS2 scan performed by the second mass spectrometer is performed during a first dead time when the polarity of the first mass spectrometer is being switched.

2. The method according to claim 1, further comprising switching the polarity of the ion source and the ion processing region of the mass spectrometer to a second polarity while the second mass spectrometer performs at least one MS1 scan or at least one MS2 scan of the ions in the one or more second ion packets with the first polarity.

3. The method according to claim 2, wherein the last MS1 scan or MS2 scan of the second scan sequence performed by the second mass spectrometer with the first polarity is at least partially performed during a second dead time when the polarity of the ion source and the ion processing region is being switched to the second polarity.

4. After switching the polarity of the ion source and the ion processing region, ionizing the sample in the ion source and the ion processing region operating with the second polarity to generate a further plurality of ions. Direct a third ion packet towards the first mass spectrometer and perform a third scan sequence including at least one MS1 scan or at least one MS2 scan of the ions in the third ion packet by the first mass spectrometer operating in the second polarity, the method according to claim 2 or 3 further comprising.

5. After performing at least one MS1 scan or at least one MS2 scan of the ions in the one or more second ion packets with the second mass spectrometer in the first polarity, switching the polarity of the second mass spectrometer to the second polarity, and at least a part of the at least one MS1 scan or at least a part of the at least one MS2 scan is performed during a first dead time in which the polarity of the first mass spectrometer is switched, the present invention Direct one or more fourth ion packets towards the second mass spectrometer and perform a fourth scan sequence including at least one MS1 scan or at least one MS2 scan of the ions in the one or more fourth ion packets by the second mass spectrometer in the second polarity, the method according to claim 4 further comprising.

6. The method according to claim 5, wherein performing at least one MS1 scan or at least one MS2 scan of the ions in the third ion packet by the first mass spectrometer operating in the second polarity is at least partially performed during a third dead time in which the polarity of the second mass spectrometer is switched to the second polarity.

7. The method according to claim 1 or 2, wherein performing at least one MS1 scan or at least one MS2 scan of the ions in the first ion packet by the first mass spectrometer operating in the first polarity is at least partially performed during a fourth dead time in which the polarity of the second mass spectrometer is switched to the first polarity.

8. The third scan sequence performed by the first mass spectrometer in the second polarity is an MS1 scan, and the method Direct the third ion packet towards the first mass analyzer, and after starting the third scanning sequence, without switching the polarity of the second mass analyzer, direct one or more fourth ion packets towards the second mass analyzer, and further include performing one or more MS2 scans of the ions in the one or more fourth ion packets with the first polarity by the second mass analyzer, wherein at least a part of the one or more MS2 scans is performed while the first mass analyzer is performing the MS1 scan. The method according to claim 4.

9. After performing the MS1 scan during the third scanning sequence with the second polarity by the first mass analyzer, switching the polarity of the ion source and the ion processing region to the second polarity; After switching the polarity of the ion source and the ion processing region to the second polarity, direct one or more fifth ion packets towards the first mass analyzer, and perform a fifth scanning sequence including one or more MS2 scans of the ions in the one or more fifth ion packets with the second polarity by the first mass analyzer. The method according to claim 8, further comprising.

10. The method according to claim 8, wherein the second mass analyzer does not have its polarity switched and only performs MS2 scans with the first polarity.

11. The method according to claim 1 or 2, wherein the method is performed within a period based on the width of the chromatographic peak of the sample when the sample elutes from the chromatographic system.

12. The method according to claim 1 or 2, wherein the first scanning sequence includes one or more MS1 scans, and the second scanning sequence includes one or more MS2 scans.

13. When MS2 scans are performed in a scanning sequence, the scanning sequence includes performing a plurality of MS2 scans respectively for a series of ion packets sequentially directed towards each of the analyzers. The method according to claim 1 or 2.

14. The method according to claim 1 or 2, wherein the first mass analyzer is an orbitrap mass analyzer, and the second mass analyzer is an ion trap mass analyzer or a time-of-flight mass analyzer.

15. The dual analyzer mass spectrometer includes one or more ion traps, and the method includes At least one of the one or more ion traps aggregates ions ionized by the ion source and the ion processing region to form an ion packet, directing the ion packet to the first mass analyzer and / or the second mass analyzer by the at least one ion trap, the method according to claim 1 or 2. **Claim 16** The method according to claim 1 or 2, wherein the first mass analyzer and / or the second mass analyzer requires a period of at least 50 ms, at least 100 ms, at least 200 ms, or at least 500 ms to switch polarities and resume mass spectrometry. **Claim 17** The method according to claim 1 or 2, wherein the mass analyzer is a High Resolution Accurate-Mass (HRAM) analyzer. **Claim 18** further ionizing the sample with the first polarity to generate a further plurality of ions, directing a further first ion packet among the plurality of ions to the first mass analyzer; performing a fifth scanning sequence including at least one MS1 scan or at least one MS2 scan of the ions in the further first packet with the first polarity by the first mass analyzer; and switching the polarity of the first mass analyzer to the second polarity after performing the at least one MS1 scan or the at least one MS2 scan, directing one or more further second ion packets among the plurality of ions to the second mass analyzer; performing a sixth scanning sequence including at least one MS1 scan or at least one MS2 scan of the ions in the one or more further second ion packets with the first polarity by the second mass analyzer; and further comprising, at least a part of the at least one MS1 scan or at least a part of the at least one MS2 scan performed by the second mass analyzer is performed during a further first dead time when the polarity of the first mass analyzer is switched, the method according to claim 1 or 2. **Claim 19** A method of operating a dual analyzer mass spectrometer in which a first mass analyzer operates in a first polarity and a second mass analyzer operates in a second polarity opposite to the first polarity to obtain MS1 and MS2 scans of cations and anions from a sample, the method comprising: Ionizing the sample in the ion source and ion processing region of the mass spectrometer to generate a plurality of ions, wherein the ion source and the ion processing region operate in a first polarity, generating the plurality of ions; Directing one or more first ion packets to the first mass analyzer and starting, by the first mass analyzer, at least one MS1 scan and / or at least one MS2 scan of the ions in the one or more first ion packets in the first polarity; After starting the at least one MS1 scan and / or the at least one MS2 scan of the one or more first ion packets by the first mass analyzer, switching the polarity of the ion source and the ion processing region to a second polarity opposite to the first polarity; After switching the polarity of the ion source and the ion processing region to the second polarity, directing one or more second ion packets to the second mass analyzer and performing, by the second mass analyzer, at least one MS1 scan and / or at least one MS2 scan of the ions in the second ion packets in the second polarity, the method comprising: At least a portion of the at least one MS1 scan and / or at least a portion of the at least one MS2 scan performed by the first mass analyzer is performed during a dead time in which the polarity of the ion source and the ion processing region is switched to the second polarity. Claim 20 The method according to claim 19, wherein the first mass analyzer performs MS1 and MS2 scans in the first polarity and the second mass analyzer performs MS1 and MS2 scans in the second polarity. Claim 21 The method according to claim 19 or 20, wherein, for the first mass analyzer and / or the second mass analyzer, the period for forming the MS1 scan is longer than the period for performing the MS2 scan. Claim 22 The one or more second ion packets include a plurality of second ion packets, and while the first mass spectrometer is performing the MS1 scan and after the polarities of the ion source and the ion processing region are switched to the second polarity, the method includes the second mass spectrometer performing one or more MS1 scans and one or more MS2 scans on the plurality of second ion packets in the second polarity. While the second mass spectrometer is performing the last scan of the scans on the plurality of second ion packets, the polarities of the ion source and the ion processing region are switched back to the first polarity, and after the polarities of the ion source and the ion processing region are switched to the first polarity, one or more third ion packets are directed to the first mass spectrometer to perform one or more MS2 scans, the method according to claim 19 or 20.

23. The method according to claim 22, wherein while the first mass spectrometer is performing the MS1 scan in the first polarity, the second mass spectrometer is performing MS1 scans and MS2 scans in the second polarity.

24. The one or more first ion packets include a first ion packet, and the first mass spectrometer performs an MS1 scan of the ions in the first ion packet in the first polarity. After switching the polarities of the ion source and the ion processing region to the second polarity and directing one or more second ion packets to the second mass spectrometer, the second mass spectrometer performs one or more MS2 scans on the one or more second ion packets in the second polarity, the method according to claim 19 or 20.

25. After performing one or more MS2 scans on the second ion packets by the second mass spectrometer, directing a third ion packet to the second mass spectrometer and performing an MS1 scan of the third ion packet in the second polarity by the second mass spectrometer. After directing the third ion packet to the second mass spectrometer to perform an MS1 scan, and while the second mass spectrometer is performing the MS1 scan of the third ion packet, switching the polarities of the ion source and the ion processing region to a second polarity opposite to the first polarity. After switching the polarities of the ion source and the ion processing region to the second polarity, directing one or more fourth ion packets toward the first mass analyzer and performing one or more MS2 scans of the ions in the one or more fourth ion packets in the first polarity by the first mass analyzer, the method according to claim 24, further comprising.

26. The method according to claim 19 or 20, wherein the method is performed within a period based on the width of a chromatographic peak of the sample when eluting from a chromatographic system.

27. The method according to claim 19 or 20, wherein the MS2 scan performed on one of the first mass analyzer and the second mass analyzer is performed in parallel with the MS1 scan performed on the other of the first mass analyzer and the second mass analyzer.

28. The method according to claim 19 or 20, wherein the first mass analyzer is an orbitrap mass analyzer and the second mass analyzer is a time-of-flight mass analyzer or an orbitrap mass analyzer.

29. The double analyzer mass spectrometer comprises one or more ion traps, and the method comprises aggregating ions ionized by the ion source and the ion processing region in at least one of the one or more ion traps to form an ion packet; directing the ion packet toward the first mass analyzer and / or the second mass analyzer by the at least one ion trap, the method according to claim 19 or 20.

30. The method according to claim 19 or 20, wherein the first mass analyzer and / or the second mass analyzer require a period of at least 50 ms, at least 100 ms, at least 200 ms, or at least 500 ms to switch the polarity and resume mass spectrometry.

31. The method according to claim 19 or 20, wherein the mass analyzer is a high-resolution accurate mass (HRAM) analyzer.

32. A mass spectrometer, comprising an ion source and an ion processing region, comprising an ionization source for generating a plurality of precursor ions from the molecules of the sample; a mass filter, an ion source and an ion processing region; An ion trap configured to aggregate or collect ions ionized by the ion source and the ion processing region to form an ion packet, and selectively direct the ion packet to a first mass spectrometer and / or a second mass spectrometer; The first mass spectrometer; A fragmentation device; The second mass spectrometer; A controller configured to cause the mass spectrometer to perform the method according to claim 1 or 19. A mass spectrometer comprising:

33. A computer program comprising computer program instructions which, when executed on a computer or controller configured to control a mass spectrometer, cause the mass spectrometer to perform the steps of the method according to claim 1 or 19. A computer program.

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