Ion trap scheme with improved mass range

Abstract

A method for trapping ions in an ion trap assembly, the method comprising: (a) Introducing ions into the ion trap arrangement, (b) applying a first RF trapping amplitude to the ion trap assembly to trap introduced ions having m / z ratios within a first range of m / z ratios; (c) cooling the trapped ions; (d) reducing the RF trapping amplitude from the first RF trapping amplitude to a second, lower RF trapping amplitude to lower the lower mass cut-off of the ion trap assembly; and (e) capturing introduced ions having m / z ratios within a second range of m / z ratios at the second, lower RF capture amplitude; wherein a lower mass limit of the second range of m / z ratios is below the lower mass cut-off of the ion trap arrangement when the first RF trapping amplitude is applied, wherein the total number of trapped ions in the ion trap assembly is kept below a threshold determined as a function of the first and second RF trapping amplitudes.

Description

Field of the invention

[0001] This invention relates to the capture of ions in an ion trap arrangement. General state of the art

[0002] Mass spectrometry is an important technique for chemical analysis. A mass spectrometer generally comprises an ion source for generating ions from a sample, various lenses, mass filters, ion trap / storage devices, and / or fragmentation devices, and one or more mass analyzers.

[0003] A critical component of a mass spectrometer is the linear ion trap. An example of such a linear ion trap is a curved linear ion trap, or C-trap, which stores / traps ions in a trapping volume using a potential well created by applying an RF potential to a set of curved, elongated rods (typically arranged as a quadrupole, hexapole, or octapole).

[0004] One application of linear ion traps is as a temporary storage device for ions prior to mass analysis. For example, a C-trap can be used to store ions and inject them into an orbital trap mass analyzer, such as the Orbitrap® device marketed by Thermo Fisher Scientific, Inc. These mass analyzers exhibit high mass accuracy and high mass resolution and are thus increasingly used for the detection of small organic molecules, such as in food and drug analysis, metabolomics, and anti-doping applications. Throughout this document, the term "mass" can be used to refer to the mass-to-charge ratio, m / z.

[0005] One of the challenges of a linear ion trap lies in the range of masses that can be trapped simultaneously. Ions are trapped in a linear trap by applying RF voltages to the longitudinal electrodes to radially confine the ions, while a static (DC) potential is applied to the end electrodes positioned at opposite axial ends of the longitudinal electrodes to axially confine the ions. The pseudopotential well created by a given applied RF voltage decreases in strength with increasing ion mass. However, higher-mass ions have similar kinetic energy to lower-mass ions. Therefore, higher-mass ions are more likely to have sufficient energy to escape the pseudopotential well created by the given applied RF trapping amplitude and thus be attenuated in the trap.Accordingly, higher-mass ions exhibit lower capture efficiencies, limiting the mass range of an ion trap. In practice, the ratio of the highest captured mass to the lowest captured mass in ion traps such as the C-trap is often limited to 15-20.

[0006] It is desirable that the mass range trapped within the linear ion trap be as broad as possible. One way to define the mass range (i.e., the range of m / z ratios) of ions in a linear ion trap is as the ratio of the highest mass to the lowest mass that can be trapped in the ion trap. For small molecule applications, a typical desired mass range might be 1200 / 15 (80), 1500 / 15 (100), or 2000 / 15 (133).

[0007] Fig. Figure 1 is a graph of RF capture amplitude (volt peak-to-peak, Vpp) applied during injection and storage versus the relative intensity of ions trapped in the linear ion trap. For this graph, the same RF amplitude was applied to the linear ion trap for both injection and storage. Each line of the graph represents a different mass of ions. It can be seen that higher-mass ions exhibit lower intensity at lower RF amplitudes.

[0008] US 6,121,610 A concerns the improvement of capture efficiency by varying the RF capture voltage during ion injection. DE 10 2005 025 497 A1 concerns a fragmentation process in which a high storage RF voltage is used for collisional fragmentation and subsequently reduced to a lower value to trap the resulting light fragment ions in the ion trap. US 2009 / 0194684 A1 concerns a process for ion fragmentation in which the RF capture voltage is reduced after the excitation of the parent ions.

[0009] The present invention seeks to increase the range of the trapping mass of an ion trap assembly, such as a linear ion trap and particularly, but not exclusively, a curved linear ion trap (C-trap). Brief description

[0010] According to a first aspect of the present invention there is provided a method of trapping ions in an ion trap assembly as described in claim 1.

[0011] The method of claim 1 comprises: (a) introducing ions into the ion trap assembly, (b) applying a first RF capture amplitude to the ion trap assembly to capture introduced ions having m / z ratios within a first range of m / z ratios, (c) cooling the captured ions, (d) reducing the RF capture amplitude from the first RF capture amplitude to a second, lower RF capture amplitude to lower the lower mass cut-off of the ion trap assembly, and (e) capturing introduced ions having m / z ratios within a second range of m / z ratios at the second, lower RF capture amplitude, wherein a lower mass limit of the second range of m / z ratios is below the lower mass cut-off of the ion trap assembly when the first RF capture amplitude is applied, wherein a total number of captured Ions in the ion trap are kept below a threshold value,which was determined as a function of the first and second RF capture amplitude.

[0012] This method improves the mass range (range of m / z ratios) of ions captured in an ion trap array. The ion trap array may include an ion trap, such as a C-trap or other linear ion trap. In a first stage of operation, higher mass ions are captured by applying a relatively higher RF trapping potential / amplitude. Although such a higher RF trapping potential / amplitude allows for the capture of relatively high m / z ions, ions below a lower mass cutoff are not captured.

[0013] By cooling the ions before reducing the RF trapping amplitude to the second, lower RF trapping amplitude, the energy of the ions within the ion trap array is dampened by collisions with inert gas molecules, and the ions relax toward the bottom of the potential well. For example, the kinetic energy of an ion entering the ion trap array is typically in the range of 1–200 eV, while the kinetic energy of such an ion after cooling is typically less than 100 meV (0.1 eV). The ions can be cooled so that they are thermalized in the trap. Accordingly, the pseudopotential well required for ion trapping after injection is higher than that required for subsequent storage of cooled ions.The cooled higher mass ions remain within the ion trap array when the RF amplitude is reduced because they do not have sufficient kinetic energy to escape the potential well created by the second, lower RF trapping amplitude.

[0014] The second, lower RF capture amplitude applied in the second stage of operation of the ion trap assembly results in a lower lower mass cutoff than that resulting from the first, higher RF capture amplitude applied in the first stage of operation of the ion trap assembly. Thus, by reducing the RF capture amplitude applied to the ion trap assembly after the higher mass ions have been cooled, it is possible to introduce and capture lower mass ions in the ion trap assembly while simultaneously retaining the higher mass ions because they have been cooled. In other words, the lower RF capture amplitude creates an RF field sufficient to retain the cooled higher mass ions within the ion trap assembly while simultaneously allowing lower mass ions to be introduced and captured within the same ion trap assembly.This, in turn, increases the usable mass range of the ion trap array compared to a method that uses a single RF amplitude or an increasing RF amplitude to introduce and trap ions. By improving the mass range of ions that can be trapped together in an ion trap array, a wider mass range of ions can be trapped in the trap compared to previous methods.

[0015] In some embodiments, step (e) comprises introducing ions into the ion trap assembly.

[0016] In some embodiments, the ion trap assembly can be configured to eject the ions from the ion trap assembly. For example, the ion trap assembly can be configured to eject the ions from the ion trap assembly to a mass analyzer that can analyze the ejected ions for their mass. The ions can thus be analyzed in a mass scan by the analyzer. A common scan type is a "full scan," which can be used as a survey scan and should cover the widest possible mass range. The invention enables an increase in the usable mass range compared to previous methods and can thus improve one of the fundamental limitations of full scans in which the ions were trapped.

[0017] In some embodiments, the method may comprise applying n further RF capture amplitudes, each intermediate between the first and second RF capture amplitudes, to the ion trap assembly, where n≥1, wherein each of the n further RF capture amplitudes causes introduced ions having a respective nth range of m / z ratios, each having lower mass limits, to be captured; wherein the method further comprises cooling the introduced ions that are captured at a relatively higher RF capture amplitude before reducing the RF capture amplitude to a relatively lower capture amplitude. This may be desirable because the change in RF capture amplitude made each time may be smaller.

[0018] In some embodiments, ions within a selected range of m / z ratios can be introduced into the ion trap device from an upstream ion device, wherein the upstream ion device transfers ions within a selected range of m / z ratios. The method further comprises adjusting the upstream ion device to reduce a lower mass limit of the selected range of m / z ratios and reducing the RF capture amplitude from the first RF capture amplitude to the second, lower RF capture amplitude synchronously or approximately synchronously with the reduction of the lower mass limit of the selected range of m / z ratios of the upstream ion device.

[0019] By adjusting the mass transfer of an upstream ion device, it is possible to select ions within the first mass range before the ion trap array captures the ions within the first mass range. Therefore, the efficiency of the mass spectrometer can be improved.

[0020] The upstream ion device can transfer ions within the first range of m / z ratios during step (a) of claim 1, such that the introduced ions of step (a) have m / z ratios within the first range of m / z ratios. The upstream ion device can transfer ions within the second range of m / z ratios during step (e) of claim 1, such that step (e) further comprises introducing ions having m / z ratios within the second range of m / z ratios into the ion trap assembly.

[0021] While the method according to claim 1 adjusts the lower mass cutoff of the ion trap assembly by adjusting the RF capture amplitude, it is also possible to adjust the lower mass cutoff of the ion trap assembly by adjusting the RF capture frequency. According to a second aspect of the present invention, a method for trapping ions in an ion trap assembly is therefore provided, as described in claim 9.

[0022] The method of claim 9 comprises (a) introducing ions into the ion trap assembly; (b) applying a first RF trapping frequency to the ion trap assembly to trap introduced ions having m / z ratios within a first range of m / z ratios; (c) cooling the trapped ions; (d) increasing the RF trapping frequency from the first RF trapping frequency to a second RF trapping frequency to lower the lower mass cut-off of the ion trap assembly; and (e) capturing introduced ions having m / z ratios within a second range of m / z ratios at the second RF capture frequency; wherein a lower mass limit of the second range of m / z ratios is below the lower mass cut-off of the ion trap assembly when the first RF capture frequency is applied, wherein a total number of trapped ions in the ion trap assembly is maintained below a threshold determined as a function of the first and second RF capture frequencies.

[0023] Product ions generated by collision-induced dissociation typically exhibit additional kinetic energy. Furthermore, the mass range of product ions generated from a precursor is broad, typically 100 - (mz), where m is the mass of the precursor ion and z is the charge of the precursor ion (and (mz) is the product of the mass of the precursor ion and the charge of the precursor ion).

[0024] Therefore, it is also desirable to improve the mass range (range of m / z ratios) of product ions that can be generated and trapped in an ion trap arrangement.

[0025] According to a third aspect of the present invention, there is provided a method for trapping product ions in an ion trap assembly configured to fragment ions, as described in claim 10. The method according to claim 10 is advantageous because it improves the mass range of product ions that can be generated and trapped in an ion trap assembly, such as a fragmentation cell, a C-trap, or other ion trap.

[0026] The method of claim 10 comprises (a) introducing precursor ions into the ion trap assembly, (b) fragmenting the introduced precursor ions to generate product ions, (c) applying a first RF capture amplitude to the ion trap assembly to capture product ions having m / z ratios within a first range of m / z ratios, (d) cooling the captured product ions, (e) reducing the RF capture amplitude from the first RF capture amplitude to a second, lower RF capture amplitude to lower the lower mass cut-off of the ion trap assembly, and (f) capturing product ions having m / z ratios within a second range of m / z ratios at the second, lower RF capture amplitude, wherein a lower mass limit of the second range of m / z ratios is below the lower mass cut-off of the ion trap assembly when the first RF capture amplitude is applied,wherein the total number of trapped product ions in the ion trap arrangement is kept below a threshold determined as a function of the first and second RF trap amplitudes.

[0027] The product ions are generated from precursor ions introduced into the ion trap array. This can occur continuously or intermittently. At a first stage of operation, higher mass product ions are captured by applying a relatively higher RF trapping potential / amplitude. Although such a higher RF trapping potential / amplitude allows for the capture of ions with relatively high m / z, ions below a lower mass cutoff are not captured.

[0028] By cooling the ions before reducing the RF capture amplitude to the second, lower RF capture amplitude, the energy of the product ions within the ion trap array is dampened by collisions with inert gas molecules, and the ions relax toward the bottom of the potential well. The product ions can be cooled so that they are thermalized within the ion trap array. Accordingly, the pseudopotential well required for capturing product ions after fragmentation is higher than that required for subsequently storing cooled product ions. The cooled, higher-mass product ions remain within the ion trap array when the RF capture amplitude is reduced because they lack sufficient kinetic energy to escape the potential well created by the second, lower RF capture amplitude.

[0029] The second, lower RF capture amplitude applied in the second stage of operation of the ion trap assembly results in a lower lower mass cutoff than that resulting from the first, higher RF capture amplitude applied in the first stage of operation of the ion trap assembly. Thus, by reducing the RF capture amplitude applied to the ion trap assembly after the higher mass ions have been cooled, it is possible to capture lower mass ions in the ion trap assembly while simultaneously retaining the higher mass ions because they have been cooled. In other words, the lower RF capture amplitude creates an RF field sufficient to retain the cooled higher mass ions within the ion trap assembly while simultaneously allowing lower mass ions to be trapped within the same ion trap assembly.This in turn increases the usable mass range of the ion trap array compared to a method that uses a single RF amplitude or an increasing RF amplitude to generate and trap product ions.

[0030] According to a fourth aspect of the present invention, there is provided a controller for controlling the trapping of ions in an ion trap assembly as described in claim 19.

[0031] According to a fifth aspect of the present invention, there is provided a further controller for controlling the trapping of ions in an ion trap assembly as described in claim 24.

[0032] According to a sixth aspect of the present invention, there is provided a further controller for controlling the fragmentation and trapping of ions in an ion trap assembly as described in claim 25.

[0033] According to a seventh aspect of the present invention, there is provided an ion trap assembly as described in claim 26.

[0034] According to an eighth aspect of the present invention, there is provided a mass spectrometer as described in claim 27. Short description of the drawings

[0035] The invention may be put into practice in many different ways and some specific embodiments will now be described by way of example only and with reference to the accompanying drawings, in which: Fig. Figure 1 is a graph of relative intensity of ions within an ion trap vs. RF amplitude applied to the ion trap for injection and storage according to known methods. Fig. Figure 2 is a schematic diagram of a first embodiment of a mass spectrometer with an ion trap according to the present invention. Fig. 3 is a flowchart illustrating a method for operating the mass spectrometer of Fig. 2 according to a first embodiment of the present invention. Fig. Figure 4 is a schematic diagram of an RF capture amplitude applied to the ion trap vs. a time diagram for the Fig. 3 described procedures. Fig. Figure 5 shows a flow chart illustrating a method for operating the mass spectrometer of Fig. 2 according to a second embodiment of the present invention. Fig. Figure 6 is a schematic diagram of an RF capture amplitude applied to the ion trap vs. a time diagram for the Fig. 5 described procedures. Fig. Figure 7 is a graph of relative intensity of ions vs. a second RF trapping amplitude applied to the ion trap according to the first embodiment of the present invention. Fig. Figure 8 is a graph of an observed trapped mass range (MR) vs. an automatic gain control (AGC) setpoint, i.e., a target number of trapped ions obtained using a calibration sample (Calmix). Fig. Figure 9(a) is a mass spectrum obtained using a calibration sample (Calmix) and a single RF amplitude according to state-of-the-art methods. Fig. 9(b) is a mass spectrum obtained using a calibration sample (Calmix) and various first and second RF amplitudes according to the first embodiment of the present invention. Fig. 10 is a flowchart illustrating a method of operating the mass spectrometer of Fig. 2 according to a third embodiment of the present invention. Fig. Figure 11 is a schematic diagram of an RF trapping amplitude applied to the ion trap array vs. a time diagram for the Fig. 10 described procedures. Fig. Figure 12(a) is a mass spectrum obtained when ions within first and second mass ranges are trapped in a fragmentation cell according to the method of the first embodiment, and ions are transferred from the fragmentation cell to an ion trap while applying the second RF trapping amplitude. A calibration sample (Calmix) was used. Fig. 12(b) is a mass spectrum obtained using a calibration sample (Calmix) according to the third embodiment of the present invention. Fig. 13 is a schematic diagram of a second embodiment of a mass spectrometer with an ion trap and an ion cooling device according to the present invention. Detailed description

[0036] Fig. Figure 2 shows a schematic arrangement of a mass spectrometer 10 suitable for carrying out methods according to embodiments of the present invention. The arrangement of Fig. Figure 2 schematically illustrates the configuration of the Q Exactive® Quadrupole-Orbitrap® mass spectrometer from Thermo Fisher Scientific, Inc.

[0037] The mass spectrometer 10 includes an ion source 20 that generates gas-phase ions to be analyzed. The ion source 20 is typically an electrospray ionization source at atmospheric pressure. These sample ions then enter a vacuum chamber of the mass spectrometer 10 and are directed through a capillary 25 into an S-lens 30.

[0038] The S-lens 30 is also known as the stacked-ring ion guide (SRIG) or RF lens. By applying RF amplitudes to the S-lens 30, an RF field is created that confines and focuses ions as they pass through the S-lens 30. The ions are focused into an injection flatapole 40, which injects the ions into a curved flatapole 50. The curved flatapole 50 guides (charged) ions along a curved path, while unwanted neutral molecules, such as entrained solvent molecules, are not guided along the curved path and are lost.

[0039] A TK lens 60 is located at the end remote from the curved flatapole 50. Ions pass from the curved flatapole 50 into a downstream quadrupole mass filter 70. The quadrupole mass filter 70 can be operated with a mass selection window so that the mass filter 70 extracts only the ions within a desired mass selection window containing ions with the m / z ratios of interest (i.e., a window containing the isotopes of interest). The mass filter is typically, but not necessarily, segmented and serves as a bandpass filter. In some operating modes, the quadrupole mass filter 70 can be operated in an essentially exclusively RF mode to transmit the widest possible mass range of ions. This is used, for example, when a "full scan" is desired and the mass range should be as wide as possible.

[0040] Ions then pass through a quadrupole exit lens / half-lens assembly 80, which controls the passage of ions into a transfer multipole 90. The transfer multipole 90 directs the mass-filtered ions from the quadrupole mass filter 70 into an ion trap, which is a curved trap (C-trap) 100. The C-trap 100 has an electrode assembly including longitudinally extending, curved rod electrodes supplied with RF voltages at RF trapping amplitudes, and end lenses supplied with DC voltages. The result is a potential well extending along the curved longitudinal axis of the C-trap 100. The C-trap 100 stores ions in a trapping volume by applying the RF trapping amplitude to the rod electrodes (typically a quadrupole, hexapole, or octapole). In other words, the C-trap 100 can operate in an “RF-only mode” for storing ions, i.e., there is no DC offset between the RF voltages.In some operating modes, a small DC offset could be applied to the rod electrodes. In some embodiments, the C-trap can be replaced with a rectilinear ion trap with straight, longitudinally extending electrodes. C-traps used according to embodiments of the present invention typically have an inscribed radius of 3 mm, a length of 25 to 30 mm, an ejection slit with a width of 0.8 mm and a length of 12 mm, an end aperture with a thickness of 1 mm, and an inscribed diameter of 2-2.5 mm.

[0041] Cooled ions are located in a cloud toward the bottom of the potential well and are then ejected perpendicularly from the C-trap 100 toward an orbital trapping device 110, such as the Orbitrap® mass analyzer sold by Thermo Fisher Scientific, Inc. Ions exit the C-trap 100, for example, by turning off the RF trapping voltage / amplitude and applying a DC pulse to one or more of the elongated longitudinal electrodes of the C-trap 100 to eject ions radially from the trap (for example, push-pull DC voltages can be applied to elongated electrodes on opposite sides of the trap). The ions are injected into the orbital trapping device 110 as coherent packets through an off-center injection aperture.Ions are then trapped within the orbital trap device 110 by a hyperlogarithmic electric field and subjected to orbital motion in coherent packets around an internal electrode. As will be understood by those skilled in the art, ion packets are detected by image currents, and a mass spectrum is then obtained by fast Fourier transform.

[0042] Fig. Figure 2 also shows a fragmentation cell 120, which allows MS / MS analysis of ions. The "dead-end" configuration of the fragmentation chamber 120 in Fig. 2, wherein precursor ions are ejected axially from the C-trap 100 in a first direction toward the fragmentation chamber 120 and the resulting fragment ions are returned to the C-trap 100 in the opposite direction, is described in more detail in WO 2006 / 103412 A2.

[0043] The mass spectrometer 10 is under the control of a controller 130, which is configured, for example, to set the appropriate potentials at the electrodes of the quadrupole mass filter 70 for focusing and filtering the ions, to set appropriate voltages at the electrode assembly of the ion trap 100 for capturing, storing, and ejecting the ions, to acquire the mass spectral data from the orbital trap device 110, to control the sequence of MS1 and MS2 scans, and so on. It is understood that the controller 130 may comprise a computer operable according to a computer program comprising instructions for causing the mass spectrometer 10 to perform the steps of the method according to the present invention. The controller 130 may comprise a trigger circuit for initiating the application of RF capture amplitudes to the electrode assembly of the ion trap 100.The controller 130 may include a timer for controlling the duration of each RF capture amplitude applied to the electrode array of the ion trap 100. Information regarding the mass range of ions to be captured by the ion trap 100 may be input to the controller 130.

[0044] An exemplary first embodiment of the method will now be described with reference to the Fig. 3 and Fig. 4. In the embodiment of the Fig. 3 and Fig. 4, the ion trap arrangement is the ion trap 100 of Fig. 2, which is a C-trap. However, it is understood that in other embodiments, the ion trap assembly could be, for example, the fragmentation chamber 120.

[0045] In step 401, sample molecules are ionized using the ESI source 20. Sample ions then enter the vacuum chamber of the mass spectrometer 10. The sample ions are guided through the capillary 25 to the S-lens 30 downstream of the ion source.

[0046] In step 402, the ions are selected according to a first mass range (first range of m / z ratios) (MR1). The first mass range (MR1) has a lower mass limit and an upper mass limit. The mass filter 70 is set to a wide pass mode by the controller 130. The ions are selected by an upstream ion device that transmits ions within the selected mass range. The upstream ion device can, for example, be one or more of the RF components of the mass spectrometer 10 upstream of the ion trap 100, such as the S-lens 30, the injection flatapole 40, and the curved flatapole 50.Optionally, the RF amplitude applied to one or more of the RF components is adjusted so that ions within the first mass range (MR1) pass through the S-lens 30, the injection flatapole 40, the curved flatapole 50, the quadrupole mass filter 70, the quadrupole exit lens / half-lens assembly 80, and the transfer multipole 90 to the ion trap 100, as discussed above.

[0047] In step 403, the ions within the first mass range (MR1) are introduced / injected into the ion trap 100 while a first RF amplitude (V1) is applied to the ion trap 100 by the controller 130. Ions within the first mass range (MR1) are trapped within the ion trap 100 by the potential well created by the first RF amplitude (V1). The potential well extends along the curved longitudinal axis of the ion trap, which is a C-trap 100. The first RF amplitude (V1) is relatively high, for example, 950 V. The first RF trap amplitude (V1) is calculated based on the lower mass limit of the first mass range (MR1).

[0048] In step 404, the ions trapped within the first mass range (MR1) in the ion trap 100 are cooled for a period of time. The trapped ions are cooled for a period of time sufficient for the trapped ions to reduce their kinetic energy so that they remain trapped after the RF trap amplitude is reduced. The period of time may be, for example, 6 ms. While the ions are being cooled, the controller 130 maintains the first RF trap amplitude (V1) applied to the ion trap 100. The ions cool due to collisions with inert gas within the ion trap 100 over the period of time. As a result of the cooling, the kinetic energy of the ions is dampened, and they relax toward the bottom of the potential well. Typically, the ions are substantially thermalized by cooling in the ion trap.

[0049] In step 405, the RF trapping amplitude applied to the ion trap 100 is reduced from the first RF trapping amplitude (V1) to a second, lower RF trapping amplitude (V2). By reducing the RF trapping amplitude, the potential well created in the ion trap 100 has a lower potential barrier (the energy required for an ion to escape the potential well). However, because the ions within the first mass range have been cooled and their kinetic energy has been dampened, these ions still do not have enough energy to overcome the potential barrier. Therefore, the ions within the first mass range (MR1) remain trapped within the ion trap 100 after reducing the RF trapping amplitude. However, the second, lower RF amplitude (V2) results in a lower lower mass cut-off (LMCO) of the ion trap compared to the first RF capture amplitude so that lower mass ions can be captured and stored.

[0050] In step 406, the selection of ions within the second mass range (MR2) (second range of m / z ratios) is performed by the upstream ion device that transmits ions within the selected mass range. As discussed above, the upstream ion device may, for example, be one or more of the RF components of the mass spectrometer 10 upstream of the ion trap 100, such as the S-lens 30, the injection flatapole 40, and the bent flatapole 50. Optionally, the RF amplitude applied to one or more of the RF components is reduced so that ions within the second mass range (MR2) pass through the S-lens 30, the injection flatapole 40, the bent flatapole 50, the quadrupole mass filter 70, the quadrupole exit lens / half-lens assembly 80, and through the transfer multipole 90 to the ion trap 100, as discussed above. The second mass range (MR2) has a lower mass limit and an upper mass limit.The lower mass limit of the second mass range is below the lower mass cut-off (LMCO) of the ion trap 100 when the first RF trapping amplitude (V1) is applied.

[0051] In step 407, the ions within the second mass range (MR2) are introduced / injected while the controller 130 maintains the application of the second, lower RF trapping amplitude (V2). The ions within the second mass range (MR2) are trapped within the ion trap by the potential well created by the second RF trapping amplitude (V2). Ions within the first mass range (MR1) were sufficiently cooled in step 404 that they do not have enough kinetic energy to escape the potential well created by the second RF trapping amplitude (V2). Therefore, the ions within the first mass range (MR1) remain trapped within the ion trap 100.

[0052] In step 408, the ions within the ion trap 100 are cooled for a period of time while the application of the second RF amplitude to the ion trap is maintained by the controller 130. The cooled ions are located in a cloud toward the bottom of the potential well.

[0053] In step 409, the RF trapping amplitude applied to the ion trap 100 may be turned off, and DC pulses may be applied to the ion trap 100 to cause both the ions within the first mass range (MR1) and the ions within the second mass range (MR2) to be ejected from the ion trap 100 and into the orbital trap mass analyzer 110. The ejection of ions from an ion trap is known.

[0054] In one example, the following RF amplitudes can be applied for selection and capture of ions within the first mass range (MR1) (steps 402 and 403 of Fig. 3 and Fig. 4), where the first mass range (MR1) has a lower mass limit of 155 m / z: The RF amplitude applied to the S-lens (30) can be 98 V; the RF amplitude applied to the injection flatapole 40 can be 25 V; the RF amplitude applied to the quadrupole mass filter 70 may be 44 V; and The RF amplitude applied to the ion trap (C-trap) 100 can be 950 V.

[0055] The following RF amplitudes can be applied for selection and capture of ions within the second mass range (MR2) (steps 406 and 407 of the Fig. 3 and Fig. 4), wherein the second mass range (MR2) has a lower mass limit of 40 m / z: The RF amplitude applied to the S-lens (30) may be 51 V; the RF amplitude applied to the injection flatapole 40 can be 25 V; the RF amplitude applied to the quadrupole mass filter 70 may be 44 V; and The RF amplitude applied to the ion trap (C-trap) 100 can be 400 V.

[0056] Fig. 4 is a graph of RF capture amplitude vs. time for the Fig. 3 described procedures. In Fig. 4, the first RF capture amplitude (V1) and the second RF capture amplitude (V2) are applied for the same time period. Accordingly, the injection and capture of ions within the first mass range (MR1) and the capture and injection of ions within the second mass range (MR2) occur for the same time period. Therefore, the ion intensities measured for the first mass range (MR1) are proportional to the ion intensities measured for the second mass range (MR2). Accordingly, the mass spectrum obtained by the mass analyzer 110 is not distorted. As shown in Fig. 4, the reduction of the RF capture amplitude from the first RF capture amplitude (V1) to the second RF capture amplitude (V2) is performed discontinuously, ie as a stepwise change.

[0057] While the Fig. 3 and Fig. 4 was described for two different mass ranges, it is possible to practice the invention using three, four, five, or more different mass ranges. Indeed, the method may include applying n further RF capture amplitudes to the ion trap 100, where n represents one or more. Each of the RF capture amplitudes may be between the first and second RF capture amplitudes. Each of the introduced ions having a respective nth mass range (range of m / z ratios) is captured by applying n further capture amplitudes to the ion trap 100. The controller 130 maintains the current RF capture amplitude for a period of time sufficient for the ions within the ion trap 100 to cool before the RF capture amplitude is reduced to a relatively lower capture amplitude.The trapped ions are cooled for a period of time sufficient for the trapped ions to reduce their kinetic energy so that they remain trapped after reducing the RF trapping amplitude.

[0058] For example, the first and second RF trapping amplitudes (V1, V2) may be the same as those used in the method of the first embodiment. Therefore, the mass range of the ions ultimately trapped within the ion trap 100 is the same as in the first embodiment. However, each of the n further RF trapping amplitudes may be intermediate RF trapping amplitudes, i.e., between those first and second RF trapping amplitudes (V1, V2). This arrangement is described with respect to the Fig. 5 and Fig. 6 is discussed in more detail. In this arrangement, instead of reducing the RF capture amplitude directly from the first to the second capture amplitude, the RF capture amplitude is reduced stepwise across the intermediate RF capture amplitudes. Therefore, the change in the RF capture amplitude performed at each time is smaller.

[0059] Alternatively, the n further RF capture amplitudes may be used to increase the mass range of ions ultimately trapped within the ion trap 100 compared to the method of the first embodiment. For example, one or more of the n further RF capture amplitudes may not lie between the first and second RF capture amplitudes. One or more of the n further RF capture amplitudes may be larger than the first RF capture amplitude (V1). Accordingly, by applying this larger RF capture amplitude before reducing the RF capture amplitude to the first RF capture amplitude (V1), it would be possible to capture ions with a higher mass than the upper mass limit of the first mass range (MR1). Alternatively or additionally, one or more of the n further RF capture amplitudes may be lower than the second RF capture amplitude (V2).By applying this lower RF capture amplitude, the lower mass cutoff of the ion trap 100 is lowered. Accordingly, by reducing the RF capture amplitude to this lower RF capture amplitude after applying the second RF capture voltage (V2), it would be possible to capture ions with a lower mass than the lower mass cutoff of the ion trap (100) when the second RF capture amplitude (V2) is applied.

[0060] In addition to applying RF capture amplitude(s) greater than V1 and / or less than V2, one or more RF capture amplitudes may also be applied between V1 and V2, as discussed above.

[0061] An exemplary second embodiment of the method will now be described with reference to the Fig. 5 and Fig. 6, where ions are trapped in the ion trap 100 of Fig. 2, which is a C-trap, is introduced and captured. The second embodiment of the method requires five different mass ranges and five corresponding RF capture amplitudes.

[0062] Steps 601, 602, 603 and 604 are the same as steps 401, 402, 403 and 404 of Fig. 3.

[0063] In step 605, the RF trap amplitude applied to the ion trap 100 is reduced from the first RF trap amplitude (V1) to a third, relatively lower RF trap amplitude (V3).

[0064] In step 606, ions are selected according to a third mass range (MR3) (third range of m / z ratios) with a lower mass limit and an upper mass limit. The lower mass limit of the third range of m / z ratios is below the lower mass cutoff of the ion trap 100 when the first RF capture amplitude (V1) is applied. The lower mass cutoff (LMCO) of the ion trap 100 with the third RF capture amplitude applied is lower than the lower mass cutoff of the ion trap 100 with the first RF capture amplitude (V1) applied.

[0065] In step 607, the ions within the third mass range (MR3) are introduced / injected and trapped within the ion trap while the controller 130 maintains the application of the third RF trapping amplitude (V3).

[0066] In step 608, the ions within the ion trap are cooled for a period of time while the application of the third RF trapping amplitude (V3) to the ion trap is maintained by the controller 130.

[0067] In step 609, the RF trap amplitude applied to the ion trap 100 is reduced from the third RF trap amplitude (V3) to a fourth, relatively lower RF trap amplitude (V4).

[0068] In step 610, ions are selected according to a fourth mass range (MR4) (fourth range of m / z ratios) with a lower mass limit and an upper mass limit. The lower mass limit of the fourth range of m / z ratios is below the LMCO of the ion trap 100 when the third RF capture amplitude (V3) is applied. The LMCO of the ion trap 100 when the fourth RF capture amplitude (V4) is applied is lower than the LMCO of the ion trap 100 when the third RF capture amplitude (V3) is applied.

[0069] In step 611, the ions within the fourth mass range (MR4) are introduced / injected and trapped within the ion trap while the controller 130 maintains the application of the fourth RF trapping amplitude (V4).

[0070] In step 612, the ions within the ion trap are cooled for a period of time while the application of the fourth RF trapping amplitude (V4) to the ion trap is maintained by the controller 130.

[0071] In step 613, the RF trapping amplitude applied to the ion trap is reduced from the fourth RF trapping amplitude to a fifth, relatively lower RF trapping amplitude (V5).

[0072] In step 614, ions are selected according to a fifth mass range (MR5) (fifth range of m / z ratios) with a lower mass limit and an upper mass limit. The fifth mass range (MR5) has a lower and an upper mass limit. The lower mass limit of the fifth range of m / z ratios is below the LMCO of the ion trap 100 when the fourth RF capture amplitude (V4) is applied. The LMCO of the ion trap 100 when the fifth RF capture amplitude (V5) is applied is lower than the LMCO of the ion trap 100 when the fourth RF capture amplitude (V4) is applied.

[0073] In step 615, the ions within the fifth mass range (MR5) are introduced / injected and trapped in the ion trap while the controller 130 maintains the application of the fifth RF trap amplitude (V5).

[0074] In step 616, the ions within the ion trap 100 are cooled for a period of time while the application of the fifth RF trap amplitude (V5) to the ion trap 100 is maintained by the controller 130.

[0075] In step 617, the RF trapping amplitude applied to the ion trap is reduced from the fifth RF trapping amplitude (V5) to the second, relatively lower RF trapping amplitude (V2).

[0076] In step 618, the ions are selected according to the second mass range (MR2) with a lower mass limit and an upper mass limit, analogous to step 406. The lower mass limit of the second range of m / z ratios is below the LMCO of the ion trap (100) when the fifth RF capture amplitude (V5) is applied. The LMCO of the ion trap (100) when the second RF capture amplitude (V2) is applied is lower than the LMCO of the ion trap 100 when the fifth RF capture amplitude (V5) is applied.

[0077] In step 619, the ions within the second mass range (MR2) are introduced / injected and trapped in the ion trap while the controller 130 maintains the application of the second, lower RF trap amplitude (V2).

[0078] In step 620, the ions within the ion trap 100 are cooled for a period of time while the application of the second RF trapping amplitude (V2) to the ion trap 100 is maintained by the controller 130.

[0079] In step 621, the RF trapping amplitude applied to the ion trap 100 may be turned off, and DC pulses may be applied to the ion trap 100 to cause the ions within the first, second, third, fourth, and fifth mass ranges (MR1, MR2, MR3, MR4, MR5) to be ejected from the ion trap 100 and into the orbital trap mass analyzer 110. The ejection of ions from an ion trap 100 is known.

[0080] Steps 605, 609, 613 and 617 are equivalent to step 405 of Fig. 3. By reducing the RF capture amplitude, the potential well created in the ion trap 100 exhibits a lower potential barrier. However, because the ions within the ion trap have been cooled and their kinetic energy has been dampened, these ions still do not have enough energy to overcome the potential barrier. Therefore, the ions remain trapped within the ion trap 100 after reducing the RF capture amplitude.

[0081] Steps 606, 610, 614 and 618 are equivalent to step 406 of Fig. 3. The selection of ions is performed by an upstream ion device. The upstream ion device may, for example, be one or more of the RF components of the mass spectrometer 10 upstream of the ion trap 100, such as the S-lens 30. Optionally, the RF amplitude applied to one or more of the RF components is adjusted so that ions within the selected mass range pass through the S-lens 30, the injection flatapole 40, the curved flatapole 50, the quadrupole mass filter 70, the quadrupole exit lens / half-lens assembly 80, and through the transfer multipole 90 to the ion trap 100, as discussed above.

[0082] Steps 607, 611, 615 and 619 are equivalent to step 407 of Fig. 3 similar.

[0083] Steps 608, 612, 616 and 620 are equivalent to step 404 of Fig. 3. By cooling the trapped ions before reducing the RF trapping amplitude, the cooled ions do not have sufficient kinetic energy to escape the potential well created by the relatively lower RF trapping amplitude. Therefore, the trapped ions remain trapped within the ion trap 100.

[0084] Fig. Figure 6 is a graph of RF capture amplitude vs. time for the Fig. 5 described procedures. In Fig. 6, each of the first, second, third, fourth, and fifth RF capture amplitudes (V1, V2, V3, V4, V5) is applied for the same time period. Accordingly, the injection and capture of ions within each mass range occurs for the same time period. Therefore, the ion intensities measured for each mass range are proportional to each other. As shown in Fig. As shown in Figure 6, each reduction of the RF capture amplitude is performed discontinuously. In Fig. 6, each of the third, fourth and fifth RF capture amplitudes (V3, V4, V5) is equally spaced between the first and second RF capture amplitudes (V1, V2).

[0085] As discussed above, Fig. Figure 1 is a graph of RF amplitude applied during injection and storage versus ion intensity within a C-trap using prior art methods. For this graph, the same RF amplitude was applied to a C-trap for both injection and capture of ions. Each line of the graph represents a different mass of ions. It can be seen that higher mass ions exhibit lower intensity at lower RF capture amplitudes. As explained in the "General State of the Art" section above, this is because the pseudopotential generated by a given RF voltage decreases in strength with increasing mass.

[0086] Fig. Figure 7 is a graph of a second RF capture amplitude (V2) applied to the ion trap 100 vs. an intensity of ions according to the method of the first embodiment of the present invention. For this graph, ions were introduced and trapped in the C-trap 100 at a first RF capture amplitude (V1) of 1500 V. The ions were then cooled. Subsequently, the RF capture amplitude was reduced to a specific second RF capture amplitude (V2) (as indicated on the x-axis of the graph) before ejecting the ions from the C-trap 100. Each line represents a different mass of ions. When comparing the Fig. 1 and Fig. 7, by capturing higher mass ions (such as m / z 1722) at a relatively high RF capture amplitude (1500 V) and cooling them prior to storage at a lower RF capture amplitude (such as 500 V), their intensity is greater than that achieved by capturing and storing these higher mass ions at a single RF capture amplitude (such as 500 V). Fig. Figure 7 also confirms that the pseudopotential well depth required to trap ions is indeed larger than the pseudopotential well depth required to store ions, that is, after the ions are cooled.

[0087] Fig. Figure 8 is a graph of an observed trapped mass range (MR) versus an automatic gain control (AGC) setpoint, i.e., a target number of trapped ions obtained using a calibration sample (Calmix). Calmix comprises a solution of caffeine (m / z 195), MRFA peptide (m / z 524), and Ultramark polymer (m / z 1122, 1222,... 1722). MR is the ratio of the highest (last) mass (Im) to the lowest (first) mass (fm) trapped in the ion trap. The AGC setpoint represents the space charge (ion population) within the ion trap. In the graph, the expected MR (expected MR) at the given RF settings is labeled for comparison with the experimentally observed values. The second RF trapping amplitude applied to the ion trap in all measurements was 300 V, corresponding to a first mass (fm) of 40. Fig. Figure 8 shows that for a given expected mass range above a threshold, the observed trapped mass range (MR) decreases as the ion population in the ion trap increases. Fig. Figure 8 thus shows a dependence of the trapped mass range (using multiple RFs during trapping according to the invention) on the space charge and that the dependence is different for different mass ranges (different RF settings). Since the mass ranges of the trapped ions depend on the first and second RF amplitudes and / or the first and second RF frequencies applied to the ion trap, in embodiments, a total number of trapped ions in the ion trap is kept below a threshold value determined as a function of the first and second RF trapping amplitudes and / or first and second RF trapping frequencies applied to the ion trap, which are applied, for example, as a function of a ratio of the first and second RF trapping amplitudes and / or the first and second RF trapping frequencies. Accordingly, Fig. 8, how a user might balance the competing needs of the broadest mass range against the highest signal-to-noise (S / N) ratio in the spectrum. The latter is very important for the depth of analysis and quantification. For example, if the desired mass range is 40, then the ion population can be 1×10 6 In contrast, if a mass range of only 30 is required, then the ion population can be further increased to 1.25×10 6 increased and the S / N of the trace components could be improved.

[0088] Fig. 9(a) and Fig. 9(b) demonstrate that the methods of the present invention achieve an increased usable mass range for an ion trap. Fig. Figure 9(a) shows a mass spectrum of a sample obtained using prior art methods, where ion injection and trapping are performed at a single RF trapping amplitude (300 V) using the calibration sample Calmix discussed above. Fig. Figure 9(b) shows a mass spectrum of the same sample as Fig. 9(a), acquired according to the methods of the present invention, wherein the first RF trap amplitude (V1) is 1000 V and the second RF trap amplitude (V2) is 300 V. The time period for changing the electronics to reduce the RF trap amplitude from the first RF trap amplitude (V1) to the second RF trap amplitude (V2) can be 0.5 to 2 ms. The ions were collision cooled during the time required to change the electronics to reduce the RF trap amplitude and also within a similar time frame as the time required to change the electronics (several ms). It can be seen that in Fig. 9(a) the mass spectrum includes ions up to m / z=200, ie the higher mass ions were attenuated. In Fig. In Figure 9(b), the mass spectrum includes ions up to m / z=540, meaning that higher mass ions were trapped and thus detected by the mass analyzer 110. Accordingly, the methods of the present invention increase the usable mass range of the ion trap 100.

[0089] While the Fig. 3 to 6 has been described with reference to a single ion trap, in a particularly advantageous embodiment of the invention, several ion traps and / or ion cooling devices can be used in the ion trap arrangement.

[0090] An exemplary third embodiment of the method will now be described with reference to the Fig. 10 and Fig. 11, which uses an ion cooling device (fragmentation cell 120 of Fig. 2) and an ion trap (ion trap 100 of Fig. 2, which is a C-trap). The fragmentation cell 120 includes an RF trapping device, such as an RF multipole, so that the fragmentation cell 120 can be operated according to the invention. The fragmentation cell 120 is operated at a higher pressure than the ion trap 100 and in a low fragmentation mode (low fragmentation including a no fragmentation mode).

[0091] In step 1001, sample molecules are ionized using the ESI source 20. Sample ions then enter the vacuum chamber of the mass spectrometer 10. The sample ions are guided through the capillary 25 to the S-lens 30 downstream of the ion source.

[0092] In step 1002, the ions are selected according to a first mass range (first range of m / z ratios) (MR1). The first mass range (MR1) has a lower mass limit and an upper mass limit. The mass filter 70 is set to a wide pass mode by the controller 130. The ions are selected by an upstream ion device that transmits ions within the selected mass range. The upstream ion device can, for example, be one or more of the RF components of the mass spectrometer 10 upstream of the ion trap 100, such as the S-lens 30, the injection flatapole 40, and the curved flatapole 50.Optionally, the RF amplitude applied to one or more of the RF components is adjusted so that ions within the first mass range (MR1) pass through the S-lens 30, the injection flatapole 40, the curved flatapole 50, the quadrupole mass filter 70, the quadrupole exit lens / half-lens assembly 80, and the transfer multipole 90 to the ion trap 100, as discussed above.

[0093] In step 1003, the ions within the first mass range (MR1) are introduced / injected into the ion trap 100. The introduced ions within the first mass range (MR1) pass through the ion trap 100 to the fragmentation cell 120, while the controller 130 applies the first RF capture amplitude (V1) to the fragmentation cell 120 and a first corresponding RF capture amplitude (V1) to the ion trap 100. The first corresponding RF trapping amplitude is applied by the controller 130 to the ion trap 100 such that the lower mass cutoff of the ion trap 100 is the same as the lower mass cutoff of the fragmentation cell in step 1003. The ion trap 100 acts as an ion conductor such that the ions within the first mass range (MR1) are not trapped within the ion trap 100 but are transferred through the ion trap 100 to the fragmentation cell 120.The controller 130 can control the RF capture amplitude applied to the ion trap 100 independently of the RF capture amplitude applied to the fragmentation cell 120. Alternatively, the RF capture amplitude can simultaneously control the RF capture amplitude applied to the fragmentation cell 120 and the ion trap 100. In step 1003, the ions within the first mass range (MR1) are transferred to the fragmentation cell 120 with minimal additional energy, thus preventing fragmentation of the transferred ions. In effect, the difference in the DC voltage applied to the ion trap 100 and the fragmentation cell 120 is minimized to prevent fragmentation during transfer. For example, the DC offset for the ion trap 100 can be 0 V and the DC offset for the fragmentation cell 120 can be -2 V.In step 1003, the ions within the first mass range (MR1) are trapped in the fragmentation cell 120 by the potential well created by the first RF trapping amplitude (V1) applied to the fragmentation cell 120 once they are transferred to the fragmentation cell 120. The first RF trapping amplitude (V1) is relatively high, for example, 950 V. The first RF trapping amplitude (V1) is calculated based on the lower mass limit of the first mass range (MR1).

[0094] In step 1004, the ions within the first mass range (MR1) trapped in the fragmentation cell 120 are cooled for a period of time within the fragmentation cell 120 while the first RF trapping amplitude (V1) is applied to the fragmentation cell 120. The first corresponding RF trapping amplitude may optionally be applied to the ion trap 100 during step 1004.

[0095] In step 1005, the ions within the first mass range (MR1) trapped in the fragmentation cell 120 are transferred from the fragmentation cell 120 to the ion trap 100 while the first RF capture amplitude (V1) is applied to the fragmentation cell 120 and the first corresponding RF capture amplitude is applied to the ion trap 100. The ions within the first mass range (MR1) are transferred to the ion trap 100 with minimal additional energy, thus preventing fragmentation of the transferred ions.

[0096] In step 1006, the ions within the first mass range (MR1) that were transferred back to the ion trap 100 are trapped in the ion trap 100 by the potential well created by the first corresponding RF trap amplitude applied to the ion trap 100. The first RF trap amplitude (V1) can optionally be applied to the fragmentation cell 120 during step 1006.

[0097] In step 1007, the ions within the first mass range (MR1) that have been transferred back to and trapped within the ion trap 100 are cooled within the ion trap 100 for a period of time while the first corresponding RF trapping amplitude is applied to the ion trap 100. The trapped ions are cooled for a period of time sufficient for the trapped ions to reduce their kinetic energy so that they remain trapped after the RF trapping amplitude is reduced. The period of time may be, for example, 6 ms. While the ions are being cooled, the controller 130 maintains the application of the first corresponding RF trapping amplitude (V1) to the ion trap 100. The ions cool due to collisions with inert gas within the ion trap 100 over the period of time. As a result of cooling, the kinetic energy of the ions is dampened and they relax towards the bottom of the potential well.Typically, the ions in the ion trap 100 are substantially thermalized by cooling. The first RF trapping amplitude (V1) can optionally be applied to the fragmentation cell 120 during step 1007.

[0098] In step 1008, the RF trapping amplitude applied to the ion trap 100 is reduced from the first corresponding RF trapping amplitude (V1) to a second, lower RF trapping amplitude (V2). By reducing the RF trapping amplitude, the potential well created in the ion trap 100 has a lower potential barrier (the energy required for an ion to escape the potential well). However, because the ions within the first mass range have been cooled and their kinetic energy has been dampened, these ions still do not have enough energy to overcome the potential barrier. Therefore, the ions within the first mass range (MR1) remain trapped within the ion trap 100 after reducing the RF trapping amplitude.However, the second, lower RF amplitude (V2) results in a lower lower mass cutoff (LMCO) of the ion trap 100 compared to the first RF capture amplitude, allowing lower mass ions to be captured and stored. The RF capture amplitude applied to the fragmentation cell 120 can be controlled independently of the RF capture amplitude applied to the ion trap. For example, the RF capture amplitude applied to the fragmentation cell 120 can be maintained at the first RF capture amplitude (V1) during step 1008. Alternatively, in step 1008, the RF capture amplitude applied to the fragmentation cell 120 can be reduced in synchronism with the RF capture amplitude applied to the ion trap 100. For example, in step 1008, the RF capture amplitude applied to the fragmentation cell 120 may be reduced from the first RF capture amplitude (V1) to a second corresponding RF capture amplitude.The lower mass cut-off of the fragmentation cell 120 when the second corresponding RF capture amplitude is applied to the fragmentation cell 120 is the same as the lower mass cut-off of the ion trap 100 when the second RF capture amplitude (V2) is applied to the ion trap 100.

[0099] In step 1009, the selection of ions within the second mass range (MR2) (second range of m / z ratios) is performed by the upstream ion device that transmits ions within the selected mass range. As discussed above, the upstream ion device may, for example, be one or more of the RF components of the mass spectrometer 10 upstream of the ion trap 100, such as the S-lens 30, the injection flatapole 40, and the bent flatapole 50. Optionally, the RF amplitude applied to one or more of the RF components is reduced so that ions within the second mass range (MR2) pass through the S-lens 30, the injection flatapole 40, the bent flatapole 50, the quadrupole mass filter 70, the quadrupole exit lens / half-lens assembly 80, and through the transfer multipole 90 to the ion trap 100, as discussed above.The second mass range (MR2) has a lower mass limit and an upper mass limit. The lower mass limit of the second mass range lies below the lower mass cutoff (LMCO) of the ion trap 100 when the first RF capture amplitude (V1) is applied.

[0100] In step 1010, the ions within the second mass range (MR2) are introduced / injected into the ion trap 100 while the controller 130 maintains the application of the second, lower RF trap amplitude (V2). The ions within the second mass range (MR2) are trapped in the ion trap 100 by the potential well created by the second RF trap amplitude (V2). Ions within the first mass range (MR1) were sufficiently cooled in step 1007, so they do not have enough kinetic energy to escape the potential well created by the second RF trap amplitude (V2). Therefore, the ions within the first mass range (MR1) remain trapped within the ion trap 100.

[0101] In step 1011, the ions within the ion trap 100 are cooled for a period of time while the application of the second RF amplitude to the ion trap 100 is maintained by the controller 130. The cooled ions are located in a cloud toward the bottom of the potential well. Cooling of ions within the ion trap 100 can occur through collisions between the trapped ions within the first mass range (MR1) and the trapped ions within the second mass range (MR2).

[0102] In step 1012, the RF trapping amplitude applied to the ion trap 100 may be turned off, and DC pulses may be applied to the ion trap 100 to cause both the ions within the first mass range (MR1) and the ions within the second mass range (MR2) to be ejected from the ion trap 100 and into the orbital trap mass analyzer 110. The ejection of ions from an ion trap 100 is known. Accordingly, both trapped ions within the first mass range (MR1) and trapped ions within the second mass range (MR2) may be analyzed together to generate a single mass spectrum with a mass range spanning both the first and second mass ranges (MR1, MR2).

[0103] The lower mass cut-off of an ion trap or fragmentation cell depends on the applied RF capture amplitude and the inscribed radii of the ion trap / fragmentation cell. In the embodiment of Fig. 10, the ion trap 100 and the fragmentation cell 120 have the same lower mass cut-off when the same RF trapping amplitude is applied. Therefore, in the embodiment of Fig. 10, the first corresponding RF capture amplitude and the first RF capture amplitude (V1) are equal. Analogously, in the embodiment of Fig. 10, the second corresponding RF capture amplitude and the second RF capture amplitude (V2) are the same. In an alternative embodiment, the ion trap 100 and the fragmentation cell 120 may have different inscribed radii. In such an embodiment, different RF capture amplitudes must be applied to them to achieve the same lower mass cutoff for the ion trap 100 as for the fragmentation cell 120. Accordingly, the first corresponding RF capture amplitude and the first RF capture amplitude (V1) would be different. Analogously, the second corresponding RF capture amplitude and the second RF capture amplitude (V2) would be different.

[0104] Typically, the ion trap (ion trap 100) is maintained at a lower pressure than the ion cooling device (fragmentation cell 120), e.g., at least 1 or at least 2 orders of magnitude lower. More generally, the pressure in the fragmentation cell 120 multiplied by the length of the fragmentation cell 120 is significantly higher than the pressure in the ion trap 100 multiplied by the length of the ion trap 100. This ensures efficient capture and transfer of high m / z ions, such as intact proteins or protein complexes.

[0105] Fig. Figure 11 is a graph of RF capture amplitude vs. time applied to both the fragmentation cell 120 and the ion trap 100 for the Fig. 10. As discussed above, in the embodiment of Fig. 10, the first corresponding RF capture amplitude and the first RF capture amplitude (V1) are equal. Similarly, the second corresponding RF capture amplitude and the second RF capture amplitude (V2) are equal. In Fig. 11, the first RF capture amplitude (V1) and second RF capture amplitude (V2) are applied for the same time period for the injection and capture of ions. Accordingly, the injection and capture of ions within the first mass range (MR1) and the injection and capture of ions within the second mass range (MR2) occur for the same time period. Therefore, the ion intensities measured for the first mass range (MR1) are proportional to the ion intensities measured for the second mass range (MR2). Accordingly, the mass spectrum obtained by the mass analyzer 110 is not distorted. Fig. Figure 11 also shows that the transfer of ions within the first mass range (MR1) from the fragmentation cell 120 to the ion trap 100 occurs before reducing the RF capture amplitude to the second RF capture amplitude (V2) and before trapping ions within the second mass range (MR2) in the ion trap 100. As in Fig. As shown in Figure 11, the reduction of the RF capture amplitude from the first RF capture amplitude (V1) to the second RF capture amplitude (V2) is performed discontinuously, i.e., as a stepwise change. Alternatively, the reduction of the RF capture amplitude could occur over a longer period of time, so that the reduction of the RF amplitude occurs as a gradient change rather than a stepwise change.

[0106] The embodiment of Fig. 10 and Fig. 11 can be particularly advantageous for addressing the challenges associated with performing mass analysis under high ion load conditions. High load conditions are common when measuring particularly demanding samples. In known high ion load setups, particularly in proteomics applications based on liquid chromatography-mass spectrometry, deposition of untransferred or uncaptured ions of relatively higher m / z can occur when ions are trapped in an ion trap / fragmentation cell. Deposition occurs when ions impact and leave residues on the rods and lenses of the ion trap / fragmentation cell due to space charge effects or unstable trajectories. Deposition leads to charging effects that can impair system performance.If the deposits form an insulating layer, the deposits can be charged by subsequent impinging ions. This would create field perturbations, altering ion trajectories and resulting in ion loss from the ion trap array.

[0107] In the embodiment of the Fig. 10 and Fig. 11, the ions are separated into ions within the first mass range (MR1) (ions with a relatively higher m / z) and ions within the second mass range (MR2) (ions with a relatively lower m / z) upstream of the ion trap 100 and the fragmentation device 120. Separating the ions allows for optimization of the capture and transfer conditions for the ions with a relatively higher m / z (ions within the first mass range) and ions with a relatively lower m / z (ions within the second mass range).

[0108] The ions within the first mass range (MR1) are trapped in a longer trap volume because the ions within the first mass range (MR1) are passed through the ion trap 100 to the fragmentation cell 120 and trapped within the fragmentation cell 120. By passing ions through the ion trap into the fragmentation cell, the RF and DC potentials applied to lenses upstream of the ion trap can focus the ions and accelerate them into the ion trap, thereby reducing deposition of high-mass ions on corresponding lenses. The lenses upstream of the ion trap are, for example, the S-lens 30 and the TK lens 50, or entrance lens of the ion trap itself. If the ions were instead trapped only in the relatively short volume of the ion trap, the DC potential applied to the entrance lens of the trap would actually have to slow down the ions passing through it.The ions are also trapped at a higher pressure because the fragmentation cell 120 operates at a higher pressure than the C-trap 100. Providing a longer, higher-pressure trapping volume for the ions with a relatively higher m / z improves the cooling of the ions with a relatively higher m / z and, accordingly, reduces deposition of ions with a relatively higher m / z during trapping. A longer, higher-pressure trapping volume is particularly preferred for, for example, intact proteins, which have a relatively long stopping distance. Due to the high momentum of the intact proteins, the required number of collisions to cool the ions is higher, requiring a longer trapping path / distance than for smaller species for a given fixed pressure. Therefore, the embodiment of the . Fig. 10 and Fig. 11 enables mass analysis of a wide mass range and reduces deposition of ions with a relatively higher m / z under high-load conditions. The lower pressure of the ion trap 100 compared to the fragmentation cell 120 can be advantageous because the co-trapped ions of the first and second mass ranges (MR1 and MR2) are subsequently accelerated from the ion trap 100 to the mass analyzer 110.

[0109] In the embodiment of the Fig. 10 and Fig. 11, ions within the first mass range (MR1) are transferred from the fragmentation cell 120 to the ion trap 100 for subsequent mass analysis. The transfer of the trapped ions to the ion trap 100 requires the application of additional energy to the trapped ions. Therefore, before the ions are transferred from the fragmentation cell 120 to the ion trap 100, the trapped ions within the first mass range (MR1) are cooled (step 1004). The trapped ions are cooled for a period of time sufficient for the trapped ions to reduce their kinetic energy, allowing them to remain in the ion trap assembly during the transfer of the ions to the ion trap 100. The ions within the second mass range (MR2) are not transferred along with the ions within the first mass range (MR1).Instead, the ions within the second mass range (MR2) are injected directly into the ion trap 100 after the ions within the first mass range (MR1) have been transferred to the ion trap 100, captured, and cooled. By transferring the ions within the first mass range (MR1) without also transferring the ions within the second mass range (MR2), ion loss during transfer is prevented. In effect, the ions within the first mass range (MR1) are transferred while the first RF capture amplitude is applied to the ion trap 100 and the fragmentation cell 120. The first RF capture amplitude (V1) creates a potential well with a potential barrier sufficient to prevent the ions within the first mass range (MR1) from escaping.

[0110] If the ions within the second mass range (MR2) were also simultaneously transferred to the ion trap 100 upon application of the first RF capture amplitude (V1), then the ions within the second mass range (MR2) would be lost. Indeed, ions below the lower mass cutoff of the ion trap 100 / fragmentation cell 120 would be lost when the first RF capture amplitude is applied. If the ions within the first mass range (MR1), even if cooled, were transferred along with the ions within the second mass range (MR2) to the ion trap 100 at the second RF capture amplitude (V2), then some of the ions within the first mass range (MR1) would be lost. The potential well created by the second RF trapping amplitude (V2) has a lower potential barrier (the energy required for an ion to escape from the potential well) than that created by the first RF trapping amplitude (V1).Accordingly, the ions within the first mass range (MR1) that received additional energy during transfer would have sufficient energy to escape the potential well created by the second RF trapping amplitude (V2). Therefore, a loss of some of the ions within the first mass range (MR1) may occur during transfer.

[0111] The balance of maximizing the mass range while providing a longer trapping volume with higher pressure for the relatively higher mass ions is determined by comparing the Fig. 12(a) and Fig. 12(b). Fig. Figure 12(a) shows a mass spectrum obtained when trapping ions within the fragmentation cell 120 is carried out according to the methods of Fig. 3 and Fig. 4 and the transfer of the ions within the first and second mass ranges together to the ion trap 100 are carried out while the second RF trapping amplitude (V2) is applied. Fig. Figure 12(b) shows a mass spectrum obtained when the method of Fig. 10 and Fig. 11. The mass spectra of the Fig. 12(a) and Fig. 12(b) were obtained using the calibration sample Calmix discussed above. When comparing the Fig. 12(a) and Fig. 12(b) it can be seen that in Fig. 12(a) only ions with an m / z up to 622 were analyzed. In fact, ions with an m / z greater than 622 were lost during the transfer of ions to the ion trap 100. In contrast, in Fig. 12(b) Ions from m / z 42 to at least 1422 were trapped in the same trapping arrangement and analyzed simultaneously. Fig. 12(b) that the embodiment of the Fig. 10 and Fig. 11 enables the capture of relatively higher mass ions in a longer capture volume at higher pressure, while simultaneously maximizing the analyzed mass range. Accordingly, the conditions are optimized for the capture of relatively high mass ions (ions within the first mass range) (MR1) and relatively lower mass ions (ions within the second mass range) (MR2), while simultaneously enabling the analysis of both high and low mass ions together.

[0112] While the Fig. 12(a) and Fig. 12(b) describe that the ion cooling device is the fragmentation cell 120 and the ion trap is the ion trap 100 of Fig. 2, which are adjacent to each other, the invention could equally be used with other combinations of adjacent ion trap / cooling devices. The ion cooling device can be an ion trap, such as a C-trap. The ion trap can be a first ion trap, and the ion cooling device can be a second ion trap.

[0113] While the data relating to Fig. 10 and Fig. 11 has been described as a C-trap 100, the ion trap may be an ion trap also configured to perform mass analysis. Accordingly, the mass analysis described in step 1012 may be performed in the ion trap, thereby avoiding the need to eject the trapped ions for mass analysis.

[0114] As in Fig. 2, the ion cooling device (fragmentation cell 120) is located downstream of the ion trap 100. However, the invention could equally be used with the ion trap downstream of the ion cooling device.

[0115] The introduced ions from step (a) according to claim 1 can be introduced into the ion cooling device (fragmentation cell 120), or can be introduced into the ion trap 100, or can be introduced into the ion trap 100 and then transferred to the ion cooling device (fragmentation cell 120). Similarly, the introduced ions from step (e) according to claim 1 can be introduced into the ion cooling device (fragmentation cell 120), or can be introduced into the ion trap 100, or can be introduced into the ion trap 100 and transferred to the ion cooling device (fragmentation cell 120).

[0116] The Fig. 10 and Fig. 11 describe the introduction and capture of the ions within the second mass range (MR2) into the ion trap 100. Alternatively, the ions within the second mass range (MR2) can be introduced and captured in the ion cooling device (fragmentation cell 120) and then transferred to the ion trap 100 (analogous to steps 1002-1005). In such an embodiment, once the ions within the first mass range (MR1) have been cooled in the ion trap 100 (step 1007), the RF capture amplitude applied to the fragmentation cell 120 is reduced to the second corresponding RF capture amplitude. The lower mass cut-off of the fragmentation cell 120 when the second corresponding RF capture amplitude is applied to the fragmentation cell 120 is the same as the lower mass cut-off of the ion trap 100 when the second RF capture amplitude is applied to the ion trap 100.The ions within the second mass range (MR2) can be selected upstream of the fragmentation cell 120 and the ion trap 100 (step 1009). The ions within the second mass range (MR2) are then introduced into the fragmentation cell 120 and trapped in the fragmentation cell 120 by applying the second corresponding RF trapping amplitude to the fragmentation cell 120. The ions within the second mass range (MR2) are then cooled and subsequently transferred to the ion trap 100 while the second corresponding RF trapping amplitude is applied to the fragmentation cell 120 and the second RF trapping amplitude (V2) is applied to the ion trap 100. The ions within the second mass range (MR2) are then trapped in the ion trap 100 by the second RF trapping amplitude (V2). Once trapped, the trapped ions within the first and second mass ranges are cooled together.Both the ions within the first mass range (MR1) and the second mass range (MR2) can be ejected together from the ion trap 100 into the mass analyzer 110. This process can be performed using three, four, five, or more different mass ranges. Indeed, the process can include applying n further RF capture amplitudes to the fragmentation cell 120, where n represents one or more. Each of the RF capture amplitudes can be between the first and second RF capture amplitudes. Each of the ions introduced into the fragmentation cell 120 with a respective nth mass range (range of m / z ratios) is captured by applying n further capture amplitudes to the fragmentation cell 120.The controller 130 maintains the current RF capture amplitude applied to the fragmentation cell 120 and the ion trap 100 for a period of time sufficient for the ions to cool within the fragmentation cell 120, be transferred to the ion trap 100, be trapped therein, and be cooled within the ion trap 100 before the RF capture amplitude is reduced to a relatively lower capture amplitude. The ions are cooled within the trap 100 once they have been transferred thereto for a period of time sufficient for the trapped ions to reduce their kinetic energy so that they remain trapped after the RF capture amplitude is reduced.

[0117] Each of the n further RF capture amplitudes can be intermediate RF capture amplitudes, i.e., between the first and second RF capture amplitudes (V1, V2). In this arrangement, instead of reducing the RF capture amplitude directly from the first to the second capture amplitude, the RF capture amplitude is reduced stepwise across the intermediate RF capture amplitudes. Therefore, the change in the RF capture amplitude implemented at each time is smaller.

[0118] Alternatively, the n additional RF trapping amplitudes can be used to increase the mass range of ions ultimately trapped within the ion trap compared to the method of Fig. 10 and Fig. 11. For example, one or more of the n further RF capture amplitudes may not lie between the first and second RF capture amplitudes. One or more of the n further RF capture amplitudes may be larger than the first RF capture amplitude (V1). Accordingly, by applying this larger RF capture amplitude before reducing the RF capture amplitude to the first RF capture amplitude (V1), it would be possible to capture ions with a higher mass than the upper mass limit of the first mass range (MR1). Alternatively or additionally, one or more of the n further RF capture amplitudes may be lower than the second RF capture amplitude (V2). By applying this lower RF capture amplitude, the lower mass cutoff of the ion trap array is lowered.Accordingly, by reducing the RF trapping amplitude to this lower RF trapping amplitude after applying the second RF trapping voltage (V2), it would be possible to trap ions with a lower mass than the lower mass cut-off of the ion trap array when the second RF trapping amplitude (V2) is applied.

[0119] The RF capture amplitudes applied to the ion cooling device and the ion trap can be changed synchronously, for example, when both devices are connected to the same RF power supply. Alternatively, the RF capture amplitude applied to the ion cooling device can be controlled independently of the RF capture amplitude applied to the ion trap.

[0120] It is understood that in the cases referred to in the Fig. 2, Fig. 10 and Fig. 11, the ions are introduced into the ion cooling device (fragmentation cell 120) in one direction and then transferred in the opposite direction from the ion cooling device to the ion trap 100, ie the ions change their direction.

[0121] In some other embodiments, the ions need not change direction after transfer between devices within the ion trap array. For example, in the Thermo Fisher Scientific Orbitrap™ Fusion Lumos mass spectrometer 15, shown schematically in Fig. 13, where components associated with the device of Fig. 2 are assigned the same reference numerals, the ion cooling device / ion trap is provided by the linear double-pressure ion trap 1140. In this case, the high-pressure ion trap 1120 may be the ion cooling device and the low-pressure ion trap 1110 may be the ion trap. The ions in this embodiment do not need to change their direction. The first and second RF trapping amplitudes are determined in the manner described above, for example, with reference to Fig. 10 and Fig. 11, to the ion cooling device (high-pressure ion trap 1120) and the ion trap (low-pressure ion trap 1110). The difference is that ions are introduced in one direction from the ion source 20 into the ion cooling device (high-pressure ion trap 1120) and then transferred in the same direction from the ion cooling device 1120 to the ion trap (low-pressure ion trap 1110), i.e., the ions do not change their direction. It is understood that the description of components of the Fig. 2 mass spectrometer equally for the components of the Fig. 13 with the same reference numbers.

[0122] In some embodiments, the method comprises ejecting trapped ions from the ion trap assembly and optionally transferring the ejected ions to a mass analyzer. In variations of any of the above embodiments, the ion trap and the mass analyzer may be the same device, i.e., such that no ejection occurs from the ion trap into the mass analyzer. For example, in the Fig. 13, the low-pressure ion trap 1110 is a mass analysis ion trap with a detector 1115.

[0123] In some embodiments, the ion cooling device (which could also be configured for ion fragmentation) could be a fragmentation cell 105 upstream of an ion trap, as in the Fig. 13. The fragmentation cell 105 may be the ion cooling device, the high-pressure ion trap 1120 may be the ion trap that receives ions transferred from the ion cooling device (fragmentation cell 105), and the low-pressure ion trap 1110 may be the mass analyzer that receives the trapped ions ejected from the high-pressure ion trap 1120. In this case, the ions do not need to be trapped in the ion cooling device (fragmentation cell 105), and the ion cooling device (fragmentation cell 105) can be operated in transfer mode. The first and second RF trapping amplitudes are otherwise applied to the ion cooling device and the ion trapping device in the manner described above, for example, with reference to Fig. 10 and Fig. 11, created.

[0124] In all of the described embodiments in which the RF capture amplitude is reduced from a first to a second capture amplitude, it is understood that instead (or additionally) the RF capture frequency may be increased from a first capture frequency to a second capture frequency.

[0125] It is understood that the ion trap assembly may comprise one or more additional electrode assemblies. At least one of the additional electrode assemblies may be configured to fragment ions.

[0126] It is understood that the control 130 of Fig. 2 may be configured to control the capture and fragmentation of ions according to the methods described herein. For example, the controller 130 may be configured to control the RF capture amplitudes / frequencies applied to the electrode assembly(s) of the ion trap(s) and / or the electrode assembly(s) of the fragmentation cell(s) according to the methods described herein. The controller 130 may be configured to apply RF capture amplitudes to the one or more further electrode assemblies for transferring product and / or precursor ions from the electrode assembly configured to fragment ions to the one or more further electrode assemblies before or after reducing the RF capture amplitude applied to the electrode assembly configured to fragment ions.

[0127] It is understood that the controller 130 may be configured to cause the ion trap assembly to eject the trapped ions from the ion trap assembly. The mass spectrometer may further include a mass analyzer configured to receive the ions ejected from the ion trap assembly and analyze the ejected ions for their mass.

[0128] It will be appreciated that the controller may be configured to continuously reduce the RF trapping amplitude applied to the electrode assembly while continuously cooling the trapped ions.

[0129] It is understood that the specific arrangement of the Fig. 2 is not essential for the methods described below. Indeed, other arrangements are suitable for carrying out the ion capture methods of embodiments of the present invention.

[0130] While the invention has been discussed with reference to a C-trap of a Q Exactive® hybrid quadrupole-Orbitrap® mass spectrometer, it is understood that the invention equally applies to other ion traps used with or without mass analyzers. For example, the ion trap assembly may comprise a C-trap, an ion guide, a fragmentation cell, a linear ion trap, a 3D ion trap, a magnetic trap, or an electrostatic trap. The present invention could be used for a linear ion trap (e.g., with curved or straight elongated electrodes) or even a 3D ion trap (Paul-type). The ion trap can be operated as an ion storage device without mass analysis of the ions, or it can be operated with mass analysis of the trapped ions, with the ion trap itself serving as the mass analyzer.The ion trap is preferably an RF multipole ion trap, preferably a quadrupole, hexapole, or octapole ion trap. Furthermore, in embodiments where the ion trap is configured to eject the stored ions to a mass analyzer for mass analysis, the mass analyzer need not be an orbital trap type, but may be a different type of mass analyzer, such as a time-of-flight (TOF) mass analyzer or FT-ICR mass analyzer, or another type of ion trap mass analyzer, including an electrostatic ion trap mass analyzer.

[0131] The process step for cooling the trapped ions has been described and illustrated in the figures as a separate, intentionally programmed period specifically for cooling. Cooling can instead occur during the period required to change the electronics to adjust the RF trapping amplitude / frequency. This can occur if the time required to change the electronics to adjust the RF trapping amplitude / frequency is longer than the time required to reduce the energy of trapped ions so that they remain trapped after the RF trapping amplitude / frequency is changed. In this case, the RF trapping amplitude / frequency may not be held constant while the trapped ions are cooled. Instead, the RF trapping amplitude can be reduced immediately after ions are trapped at the relatively higher RF trapping amplitude.Cooling would then occur while adjusting the electronics to decrease the RF trapping amplitude. Similarly, the RF trapping frequency can be increased after trapping ions at the relatively lower RF trapping frequency. Cooling would then occur while adjusting the electronics to increase the RF trapping frequency. Typical cooling times can be on the order of at least 1-10 ms or at least 1-5 ms, e.g., at least 1 ms, at least 2 ms, at least 3 ms, at least 4 ms, or at least 5 ms. A typical cooling time, for example, for peptides and singly charged ions in the range of 400 to 1000 Th would be a few ms at a background pressure of 1×10. -3 mbar.

[0132] The method of the present invention could be applied with steps 405 and 406 beginning either in any order or simultaneously. For example, while the RF capture amplitude is reduced from the relatively higher to the relatively lower capture amplitude, the RF amplitude applied to the other components of the mass spectrometer upstream of the ion trap can also be reduced at the same time. The same applies to the method of Fig. 5 and Fig. 6 and the procedure of Fig. 10 and Fig. 11. For example, steps 605 and 606 can be initiated either in any order or simultaneously. Similarly, steps 1008 and 1009 can be initiated either in any order or simultaneously.

[0133] In the process of Fig. 3, Fig. 5 and Fig. 10, the ions can be introduced into the ion trap assembly as a continuous ion stream while the other steps of the method are carried out. Of course, only the ions in the relevant mass range are selected by the upstream ion device when the relevant RF amplitude is applied. Similarly, only the ions in the relevant mass range are trapped in the ion trap assembly when the relevant RF trapping amplitude is applied. For example, with reference to the method of Fig. 3, the introduction of ions into the ion trap assembly can be carried out continuously while steps 402 to 408 are performed. Analogously, the introduction of ions into the ion trap assembly can be carried out continuously while steps 602 to 620 are performed. Alternatively, the introduction of ions into the ion trap assembly can be carried out intermittently. For example, ions can be introduced into the ion trap assembly only when the relevant RF capture amplitude is applied and stopped while the RF capture amplitude is reduced and / or cooling is carried out. For example, with reference to the method of Fig. 3, a first introduction of ions may be carried out while steps 402 and 403 are being executed. While steps 404 and 405 are being executed, the introduction of ions may be stopped. A second introduction of ions may be carried out while steps 406 and 407 are being executed. While steps 408 and 409 are being executed, the introduction of ions may be stopped. In general, a period of introduction of ions into the ion trap assembly is controlled so that the ion trap assembly is not overfilled, i.e., to avoid space charge effects described above with respect to Fig. 8. The time period for introducing ions into the ion trap arrangement thus typically depends on the ion current.

[0134] Each mass range (MR) may or may not overlap with others. For example, the first and second mass ranges (MR1, MR2) may overlap. The intensities of ions within the overlap range may not be proportional to the intensities of ions outside the overlap range. The controller 130 may be configured to compensate for this, ensuring that the relative abundances in the resulting mass spectrum are not distorted by double counting ions in the overlapping range(s).

[0135] The RF capture amplitude vs. time graph of the Fig. 4, Fig. 6 and Fig. Figure 9 shows that the reduction in the RF trapping amplitude occurs discontinuously. The reduction in the RF trapping amplitude can instead occur continuously. Optionally, the method can include continuously cooling the trapped ions while continuously reducing the RF trapping amplitude. By continuously reducing the RF trapping amplitude, the LMCO of the ion trap array is continuously reduced from the LMCO of the first RF trapping amplitude (V1) to the LMCO of the second, lower RF trapping amplitude (V2). The lower mass limit of the selected mass range transmitted by the upstream ion device can also be continuously reduced.For example, if the upstream ion device is one or more of the RF components upstream of the ion trap assembly, such as the S-lens 30, the lower mass limit of the selected mass range is continuously lowered by continuously reducing the RF amplitude applied to the RF component(s) upstream of the ion trap assembly. The reduction in the RF capture amplitude applied to the ion trap assembly can be continuously reduced in synchronization with the lowering of the lower mass limit of the selected mass range. Therefore, ions with decreasing m / z ratios pass through the S-lens 30, the injection flatapole 40, the curved flatapole 50, the quadrupole mass filter 70, the quadrupole exit lens / half-lens assembly 80, the transfer multipole 90, and are introduced and trapped in the ion trap assembly.The rate at which the RF capture amplitude is reduced is selected by the controller 130 so that ions with higher mass within the ion trap assembly have sufficient time to reduce their kinetic energy so that they remain trapped in the ion trap assembly after the RF capture amplitude is reduced to introduce ions with a lower mass ratio. Ions can be introduced into the ion trap assembly while the RF capture amplitude is reduced. The ions can be continuously introduced into the ion trap assembly while the RF capture amplitude is applied. Alternatively, the ions can be introduced intermittently as a series of multiple injections into the ion trap assembly to avoid overfilling the ion trap assembly. In general, a period of introduction of ions into the ion trap assembly is controlled so that the ion trap assembly is not overfilled, i.e.to avoid space charge effects mentioned above with respect to . Fig. 8. A period of introduction of ions into the ion trap assembly thus typically depends on the ion current. In the embodiment where the RF trap amplitude is continuously reduced, the rate of this reduction may be constant. Alternatively, the rate of reducing the RF trap amplitude may not be constant. For example, the rate of reducing the RF trap amplitude may be decreased as the RF trap amplitude is reduced. Alternatively, the rate of reducing the RF trap amplitude may be increased as the RF trap amplitude is reduced. Optionally, the first RF trap amplitude (V1) may be applied constantly for a certain period of time before the RF trap amplitude is continuously reduced to the second RF trap amplitude (V2). The second RF trap amplitude (V2) may then be applied constantly for a certain period of time.

[0136] The RF capture amplitude vs. time graphs of the Fig. 4, Fig. 6 and Fig. 9 show that each RF trap amplitude is applied for the same time duration. This is desirable to avoid distortions in the resulting mass spectrum, as discussed above. The RF trap amplitudes can instead be applied for different time durations. Alternatively, some of the RF trap amplitudes can be applied for the same time duration and some of the RF trap amplitudes for a different time duration. Applying different RF trap amplitudes for different time durations can result in distortions in the ion intensities. The controller 130 can be further configured to compensate for such distortions so that the peaks of the mass spectrum are proportional.

[0137] Fig. Figure 6 shows that each of the third, fourth, and fifth RF capture amplitudes (V3, V4, V5) is equally spaced between the first and second RF capture amplitudes (V1, V2). However, the third, fourth, and fifth RF capture amplitudes (V3, V4, V5) may not be equally spaced between the first and second RF capture amplitudes (V1, V2).

[0138] Embodiments of the present invention discuss the selection of ions within a specific mass range upstream of the ion trap assembly by an upstream ion device. Any ion device with an adjustable mass transfer profile (i.e., variable upper mass cutoff and / or lower mass cutoff) can be used to perform this selection. For example, the mass filter 70 can be set to the desired mass range by the controller 130, so that the mass filter 70 filters the sample ions according to the desired mass range. This option is less desirable for a wide "full MS" scan, since mass range selection using the mass filter 70 typically provides mass ranges that are too narrow.Embodiments of the present invention discuss adjusting the upstream ion device to change the lower mass limit of the selected mass range of ions transmitted by the upstream ion device. Alternatively or additionally, the upstream ion device may be adjusted to change the upper mass limit of the selected mass range of ions transmitted by the upstream ion device.

[0139] In some embodiments, ions may be selected by an upstream ion device (upstream of the ion trap), preferably a mass selector such as a mass filter 70, such that the first and second ranges of m / z ratios (and optionally n further ranges of m / z ratios) are selected, preferably non-overlapping. This may enable increased analysis dynamic range and better quantification. Thus, in some embodiments, ions in a first range of m / z ratios may be selected and introduced into the ion trap assembly while the first RF trapping amplitude is applied to the ion trap to capture the introduced ions having m / z ratios within the first range of m / z ratios and to cool the trapped ions.The RF capture amplitude is then reduced from the first RF capture amplitude to the second, lower RF capture amplitude to lower the lower mass cutoff of the ion trap array. Ions can be selected within a second range of m / z ratios (not overlapping the first range) and introduced into the ion trap array, while the second, lower RF capture amplitude is applied to the ion trap array to capture the introduced ions having m / z ratios within the second range of m / z ratios. In this case, the upper limit of the second mass range is sharply defined by the mass filter, while the lower limit could preferably be defined by the same mass filter or by the lower mass cutoff corresponding to the second RF amplitude. The m / z ratios of the second range of m / z ratios are preferably below the m / z ratios of the first range.In such embodiments, a lower mass limit, and in some cases the upper mass limit, of the second range of m / z ratios is below the lower mass cut-off of the ion trap assembly when the first RF trapping amplitude is applied.

[0140] The selection of ions according to a specific mass range upstream of the ion trap assembly is optional. For example, the ion trap assembly can capture all sample ions generated by ion source 20, and the RF capture amplitude applied to the ion trap assembly controls the ion capture of ions so that only those within the desired mass range are captured.

[0141] Before injection into the mass analyzer 110, the ions within the first mass range (MR1) and / or ions within the second mass range (MR2) can be fragmented in the fragmentation cell 120. The fragments can be accumulated in the C-trap 100 before being ejected into the mass analyzer as a single pulse for acquisition as a single spectrum. Alternatively, if the mass analyzer 110 is a TOF mass analyzer, fragment ions in the fragmentation cell 120 can continuously exit this fragmentation cell 120.

[0142] Embodiments of the present invention can be applied to the capture of product / fragment ions generated from precursor ions. The ion trap assembly, which may be an ion trap, may be configured to fragment ions, or it may comprise a device for fragmenting ions. Precursor ions may be introduced into the ion trap assembly and fragmented to generate product ions. The RF trapping amplitude applied to the ion trap assembly can be varied to capture product ions within a specific range of m / z ratios. For example, product ions with m / z ratios within a first range of m / z ratios may be captured in the ion trap assembly by applying the first RF trapping amplitude.These trapped product ions can be cooled to reduce their energy so that the trapped product ions remain trapped in the ion trap assembly after the RF trap amplitude is reduced. The RF trap amplitude can be reduced to a second, relatively lower RF trap amplitude to lower the lower mass cutoff of the ion trap. Product ions with m / z ratios within a second range of m / z ratios can be trapped at the second RF trap amplitude. The lower mass limit of the second range of m / z ratios is below a lower mass cutoff of the ion trap assembly when the first RF trap amplitude is applied.

[0143] The precursor ions can be continuously introduced into the ion trap assembly and / or fragmented in the ion trap assembly. For example, step (a) of claim 10 can be performed continuously while steps (b) to (f) are performed. Alternatively, introduction and fragmentation of precursor ions can be intermittent to avoid overfilling the ion trap assembly. For example, introduction and fragmentation of ions can only occur for a period of time while the first RF trapping amplitude is applied to capture a desired number of product ions. Introduction and fragmentation of ions can be stopped while the trapped product ions are cooled in step (d) of claim 10. Introduction and fragmentation of ions can be repeated for a period of time while the second RF trapping amplitude is applied to capture a desired number of product ions.The product ions captured by the first RF capture amplitude may be generated from the same precursor ions as the product ions captured by the second RF capture amplitude. Alternatively, the product ions captured by the first RF capture amplitude may be generated from different precursor ions than the product ions captured by the second RF capture amplitude. The product ions captured by the first RF capture amplitude may be generated with the same or a different collision energy than the product ions captured by the second RF capture amplitude. The product ions captured by the first RF capture amplitude and the product ions captured by the second RF capture amplitude may be ejected together into a mass analyzer.

[0144] The method for trapping ions in an ion trap assembly configured to fragment ions may comprise applying n further RF trap amplitudes, each intermediate between the first and second RF trap amplitudes, to the ion trap assembly, where n≥1, wherein each of the n further RF trap amplitudes causes product ions having a respective nth range of m / z ratios, each having lower mass limits, to be trapped; the method further comprising cooling the product ions that are trapped at a relatively higher RF trap amplitude before reducing the RF trap amplitude to a relatively lower trap amplitude.

[0145] The method for capturing product ions in an ion trap configured to fragment ions can, in some embodiments, be applied to the fragmentation cell 120 of the illustrated mass spectrometer. In some embodiments, the fragmentation cell 120 includes an RF trapping device, such as an RF multipole, so that the fragmentation cell 120 can be operated according to the invention. The fragmentation cell 120 is operated at a higher pressure than the ion trap 100 and can be operated in high and low fragmentation modes (low fragmentation including a no fragmentation mode), for example, by applying appropriate voltage offsets between the ion trap 100 and the fragmentation cell 120.In some methods of operation, after capturing product / fragment ions generated from precursor ions in the fragmentation cell 120 according to the invention, the product / fragment ions can be transferred from the fragmentation cell 120 to the ion trap 100 while the second, lower RF capture amplitude is applied. Accordingly, in some embodiments, captured product / fragment ions with m / z ratios within the first and second ranges of m / z ratios (MR1, MR2) are transferred from the ion trap configured to fragment ions to another ion trap at a different pressure than the ion trap configured to fragment ions, and the ions are transferred while the second, lower RF capture amplitude (V2) is applied. Typically, the another ion trap is maintained at a lower pressure than the ion trap configured to fragment ions, e.g.a pressure at least 1 or at least 2 orders of magnitude lower.

[0146] More generally, the pressure in the ion trap configured for fragmentation, multiplied by the length of the ion trap configured for fragmentation, is significantly higher than the pressure of the other ion trap, multiplied by the length of the other ion trap. This ensures efficient capture and transfer of high m / z ions, such as intact proteins or protein complexes.

[0147] The above-described embodiments, which include fragmentation of the ions in the ion trap, can also be applied mutatis mutandis to an ion trap arrangement comprising a plurality of electrode arrangements, wherein at least one of the electrode arrangements is configured to fragment ions. For example, the above-described embodiments, which include fragmentation of the ions in the ion trap, can also be applied mutatis mutandis to the arrangement described with reference to Fig. 10 and Fig.11, wherein the ion trap assembly to which the RF capture amplitudes are applied comprises the ion cooling device (fragmentation cell 120) and the ion trap (ion trap 100). For example, the fragmentation steps may be applied to the ions introduced into the fragmentation cell 120 when the first and second RF capture amplitudes are applied. In this way, product ions having m / z ratios within a first range of m / z ratios (MR1) may be captured along with product ions having m / z ratios within a second range of m / z ratios (MR2). The lower mass limit of the second range of m / z ratios (MR2) may be below the lower mass cutoff of the ion trap assembly when the first RF capture amplitude (V1) is applied.

[0148] For example, precursor ions can be introduced into the ion cooling device (fragmentation cell 120) of the ion trap assembly. The precursor ions can then be fragmented in the ion cooling device to generate product ions. The product ions with m / z ratios within the first range of m / z ratios can be captured and cooled in the ion cooling device while the first RF capture amplitude (V1) is applied to the ion cooling device. The product ions can then be transferred from the ion cooling device to the ion trap (ion trap 100) with minimal additional energy while the first corresponding RF capture amplitude is applied to the ion trap and the first RF capture amplitude (V1) is applied to the ion cooling device. Accordingly, during the transfer, the lower mass cutoff of the ion trap is the same as the lower mass cutoff of the ion cooling device.The product ions can then be captured and cooled in the ion trap while the first corresponding RF capture amplitude is applied. Once cooled, the RF capture amplitude applied to the ion cooling device is reduced to the second corresponding RF capture amplitude. Optionally, the RF capture amplitude applied to the ion trap is reduced to the second RF capture amplitude (V2) so that the lower mass cutoff of the ion trap is the same as the lower mass cutoff of the ion cooling device. Further fragmentation of precursor ions can optionally take place in the ion cooling device. The product ions with m / z ratios within the second range of m / z ratios can then be captured in the ion cooling device by applying the second corresponding RF capture amplitude to the ion cooling device.The product ions can be cooled in the ion cooling device while the second corresponding RF capture amplitude is applied to the ion cooling device. The product ions can then be transferred from the ion cooling device to the ion trap (ion trap 100) with minimal additional energy while the second corresponding RF capture amplitude is applied to the ion cooling device and the second RF capture amplitude (V2) is applied to the ion trap. The product ions can then be trapped in the ion trap by applying the second RF capture amplitude (V2) to the ion trap. In such embodiments, it is preferred that the ion cooling device be located upstream of the ion trap.Subsequently, the product ions with m / z ratios within the first and second ranges of m / z ratios (MR1, MR2) can be transferred to another ion trap with a different pressure than the ion trap of the ion trap array while the second RF capture amplitude (V2) is applied. Controller 130 can be configured to control the capture and fragmentation of ions according to such a process.

[0149] Optionally, the additional ion trap can have a lower pressure than the ion trap and the ion cooling device of the ion trap assembly. The pressure of the additional ion trap, multiplied by the length of the additional ion trap, can be lower than the pressure of the ion trap of the ion trap assembly, multiplied by the length of the ion trap of the ion trap assembly.

[0150] The product ions within the second mass range (MR2) may not be captured in the ion cooling device and transferred to the ion trap. Instead, the product ions within the second mass range (MR2) may pass through the ion cooling device to the ion trap while the second RF capture amplitude is applied to the ion cooling device and to the ion trap. The product ions within the second mass range (MR2) may then be captured in the ion trap by applying the second RF capture amplitude (V2) to the ion trap once the product ions within the first mass range (MR1) have been cooled. In an alternative embodiment, fragmentation may be performed in the ion trap of the ion trap assembly instead of the ion cooling device. The product ions may then be transferred to the ion cooling device and captured therein.Alternatively, both the ion trap and the ion cooling device can fragment the precursor ions to generate product ions. The first RF capture amplitude (V1) can be applied to the ion trap to capture product ions within the first mass range. The second RF capture amplitude (V2) can be applied to the ion cooling device to capture product ions within the second mass range. The trapped ions can be cooled before being transferred to the further ion trap.

[0151] The invention has been described with respect to lowering the lower mass cutoff of the ion trap assembly by reducing the RF capture amplitude. However, it is also possible to lower the lower mass cutoff of the ion trap assembly by increasing the RF capture frequency applied to the ion trap assembly. Furthermore, it is possible to change both the RF capture amplitude and the RF capture frequency, so that the net effect is to lower the lower mass cutoff of the ion trap assembly.

[0152] Indeed, the method could be considered as a method for trapping ions in an ion trap assembly, the method comprising: introducing ions into the ion trap assembly, applying a first RF trapping waveform to the ion trap assembly for trapping introduced ions having m / z ratios within a first range of m / z ratios, cooling the trapped ions, changing the RF trapping waveform from the first RF trapping waveform to a second RF trapping waveform for lowering the lower mass cutoff of the ion trap assembly; and trapping the introduced ions having m / z ratios within a second range of m / z ratios at the second RF trapping waveform, wherein a lower mass limit of the second range of m / z ratios is below the lower mass cutoff of the ion trap assembly when the first RF trapping waveform is applied.

[0153] The first RF capture waveform may comprise a first RF capture amplitude, and the second RF capture waveform may comprise a second RF capture amplitude, wherein the first RF capture amplitude is greater than the second RF capture amplitude.

[0154] The first RF capture waveform may comprise a first RF capture frequency, and the second RF capture waveform may comprise a second RF capture frequency, wherein the first RF capture frequency is less than the second RF capture frequency.

Claims

[1] A method for trapping ions in an ion trap assembly, the method comprising: (a) Introducing ions into the ion trap arrangement, (b) applying a first RF trapping amplitude to the ion trap assembly to trap introduced ions having m / z ratios within a first range of m / z ratios; (c) cooling the trapped ions; (d) reducing the RF trapping amplitude from the first RF trapping amplitude to a second, lower RF trapping amplitude to lower the lower mass cut-off of the ion trap assembly; and (e) capturing introduced ions having m / z ratios within a second range of m / z ratios at the second, lower RF capture amplitude; wherein a lower mass limit of the second range of m / z ratios is below the lower mass cut-off of the ion trap arrangement when the first RF trapping amplitude is applied, wherein the total number of trapped ions in the ion trap assembly is kept below a threshold determined as a function of the first and second RF trapping amplitudes. [2] Method according to one of the preceding claims, further comprising: applying n further RF trapping amplitudes, each lying between the first and second RF trapping amplitudes, to the ion trap assembly, where n≥1, wherein each of the n further RF trapping amplitudes causes introduced ions having a respective n-th range of m / z ratios, each having lower mass cutoffs, to be trapped; the method further comprising cooling the introduced ions trapped at a relatively higher RF trapping amplitude before reducing the RF trapping amplitude to a relatively lower trapping amplitude. [3] The method of claim 2, wherein the first, the second and / or each of the n further intermediate RF capture amplitudes are applied for the same time period, or wherein at least some of the first, the second and / or each of the n further intermediate RF capture amplitudes are applied for different times. [4] A method according to any one of claims 1 to 3, wherein the ion trap assembly comprises an ion trap and an ion cooling device. [5] The method of claim 4, wherein in step (b) the first RF trapping amplitude is applied to the ion cooling device such that ions having m / z ratios within the first range of m / z ratios are trapped in the ion cooling device; wherein in step (c) the trapped ions are cooled in the ion cooling device; wherein the method comprises step (c)(i) after step (c) and before step (d) comprising transferring the trapped ions from the ion cooling device to the ion trap while applying the first RF trapping amplitude to the ion cooling device and while applying a corresponding first RF trapping amplitude to the ion trap such that the ion trap and the ion cooling device have the same lower mass cut-off during the ion transfer in step (c)(i), and trapping the transferred ions in the ion trap at the corresponding RF trapping amplitude; wherein in step (d) the RF capture amplitude applied to the ion trap is reduced to the second, lower RF capture amplitude to lower the lower mass cut-off of the ion trap, preferably wherein in step (d) the RF capture amplitude applied to the ion cooling device is reduced to a corresponding second RF capture amplitude such that the ion trap and the ion cooling device have the same mass cut-off during step (d). [6] The method of claim 5, wherein the method comprises applying n further RF trapping amplitudes, each intermediate between the first RF trapping amplitude and the second corresponding RF trapping amplitude, to the ion cooling device, where n≥1, wherein each of the n further RF trapping amplitudes causes introduced ions having a respective n-th range of m / z ratios, each having lower mass limits, to be trapped in the ion cooling device;the method further comprising: cooling the introduced ions trapped in the ion cooling device at a relatively higher RF trapping amplitude, transferring the trapped ions to the ion trap while applying the relatively higher RF trapping amplitude, trapping the transferred ions in the ion trap by applying the relatively higher RF trapping amplitude, and cooling the trapped ions in the ion trap before reducing the RF trapping amplitude to a relatively lower trapping amplitude; [7] A method according to any one of claims 4 to 6, wherein the introduced ions of step (a) are introduced into the ion cooling device. [8] A method according to any one of claims 4 to 7, wherein step (e) comprises introducing ions into the ion trap assembly, wherein the introduced ions from step (e) are introduced into the ion trap. [9] A method for trapping ions in an ion trap assembly, the method comprising: (a) introducing ions into the ion trap array; (b) applying a first RF trapping frequency to the ion trap assembly to trap introduced ions having m / z ratios within a first range of m / z ratios; (c) cooling the trapped ions; (d) increasing the RF trapping frequency from the first RF trapping frequency to a second RF trapping frequency to lower the lower mass cut-off of the ion trap assembly; and (e) capturing introduced ions having m / z ratios within a second range of m / z ratios at the second RF capture frequency; wherein a lower mass limit of the second range of m / z ratios is below the lower mass cut-off of the ion trap arrangement when the first RF trapping frequency is applied, wherein the total number of trapped ions in the ion trap assembly is kept below a threshold determined as a function of the first and second RF trapping frequencies. [10] A method for trapping ions in an ion trap assembly, the ion trap assembly being configured to fragment ions, the method comprising: (a) Introducing precursor ions into the ion trap arrangement, (b) fragmenting the introduced precursor ions to generate product ions; (c) applying a first RF trapping amplitude to the ion trap assembly to trap product ions having m / z ratios within a first range of m / z ratios; (d) cooling the captured products; (e) reducing the RF trapping amplitude from the first RF trapping amplitude to a second, lower RF trapping amplitude to lower the lower mass cut-off of the ion trap assembly; and (f) capturing product ions with m / z ratios within a second range of m / z ratios at the second, lower RF capture amplitude; wherein a lower mass limit of the second range of m / z ratios is below the lower mass cut-off of the ion trap arrangement when the first RF trapping amplitude is applied, wherein the total number of trapped product ions in the ion trap arrangement is kept below a threshold determined as a function of the first and second RF trap amplitudes. [11] A method according to any one of the preceding claims, wherein the ion trap arrangement is an ion trap. [12] The method of claim 11, wherein the method further comprises: (g) transferring trapped product ions of the first and second range of m / z ratios to a further ion trap, wherein the further ion trap has a different pressure than the ion trap configured to fragment ions, preferably wherein the further ion trap is maintained at a lower pressure than the ion trap configured to fragment ions, wherein the ions are further transferred to the further ion trap while the second, lower RF trapping amplitude is applied. [13] The method of claim 10, wherein the ion trap assembly comprises an ion trap and an ion cooling device, wherein the ion trap and / or the ion cooling device are configured to fragment ions. [14] The method of claim 13, wherein in step (c) the first RF trapping amplitude is applied to the ion cooling device such that the product ions having m / z ratios within the first range of m / z ratios are trapped in the ion cooling device; wherein in step (d) the trapped ions are cooled in the ion cooling device; wherein the method after step (d) and before step (e) comprises step (d)(i) which comprises transferring the trapped ions from the ion cooling device to the ion trap while applying the first RF trapping amplitude to the ion cooling device and while applying a first corresponding RF trapping amplitude to the ion trap such that the ion trap and the ion cooling device have the same lower mass cut-off during the ion transfer in step (d)(i), and trapping the transferred ions in the ion trap at the first corresponding RF trapping amplitude; wherein in step (e) the RF capture amplitude applied to the ion trap is reduced to the second, lower RF capture amplitude to lower the lower mass cut-off of the ion trap. [15] The method of claim 14, wherein in step (e) the RF trapping amplitude applied to the ion cooling device is reduced to a corresponding second RF trapping amplitude such that the ion trap and the ion cooling device have the same lower mass cut-off during step (e); wherein in step (f) the ions within the range of m / z ratios are trapped in the ion cooling device by the corresponding second RF trapping amplitude applied to the ion cooling device; the method further comprising step (f)(i) which comprises transferring the trapped ions within the second mass range from the ion cooling device to the ion trap while applying the corresponding second RF trapping amplitude to the ion cooling device and the second RF trapping amplitude to the ion trap, and trapping the transferred ions with m / z ratios within the second range of m / z ratios in the ion trap at the second RF trapping amplitude. [16] The method of claim 15, wherein the method comprises applying n further RF trapping amplitudes, each intermediate between the first RF trapping amplitude and the second corresponding RF trapping amplitude, to the ion cooling device, where n≥1, wherein each of the n further RF trapping amplitudes causes introduced ions having a respective n-th range of m / z ratios, each having lower mass limits, to be trapped in the ion cooling device;the method further comprising: cooling the introduced ions trapped in the ion cooling device at a relatively higher RF trapping amplitude, transferring the trapped ions to the ion trap while applying the relatively higher RF trapping amplitude, trapping the transferred ions in the ion trap by applying the relatively higher RF trapping amplitude, and cooling the trapped ions in the ion trap before reducing the RF trapping amplitude to a relatively lower trapping amplitude; [17] Method according to one of claims 4 to 7 or 13 to 16, wherein the ion cooling device has a different pressure than the ion trap, preferably wherein the ion cooling device has a higher pressure than the ion trap. [18] A method according to any preceding claim, wherein the lower mass limit of the first range of m / z ratios is greater than an upper mass limit of the second range of m / z ratios such that the first range of m / z ratios and the second range of m / z ratios do not overlap. [19] A controller for controlling the capture of ions in an ion trap assembly, the ion trap assembly comprising an electrode assembly, the controller being configured to: Causing ions to be introduced into the ion trap assembly; Applying a first RF trapping amplitude to the electrode assembly to trap introduced ions having m / z ratios within a first range of m / z ratios for a duration sufficient to allow cooling of the trapped introduced ions; Reducing the RF capture amplitude applied to the electrode assembly from the first RF capture amplitude to a second, lower RF capture amplitude that captures introduced ions with m / z ratios within a second range of m / z ratios, wherein a lower mass limit of the second range of m / z ratios is below the lower mass cut-off of the ion trap arrangement when the first RF trapping amplitude is applied, wherein the controller is configured to determine a threshold value for a total number of trapped ions within the ion trap assembly as a function of the first and second RF trap amplitudes, wherein the controller is further configured to control the application of the first and second RF trapping amplitudes such that a total number of trapped ions in the ion trap assembly is maintained below the threshold. [20] The controller of claim 19, wherein the ion trap assembly is an ion trap. [21] A controller according to claim 19 or 20, wherein the controller is further configured to apply n further RF trapping amplitudes, each intermediate the first and second RF trapping amplitudes, to the ion trap assembly, where n≥1, each of the n further RF trapping amplitudes causing introduced ions to be trapped having a respective n-th range of m / z ratios, each having lower mass limits, the or each of the n further RF trapping amplitudes being applied for a duration sufficient to allow cooling of the ions trapped at that n-th RF trapping amplitude. [22] The controller of claim 19, wherein the ion trap assembly comprises an ion trap and an ion cooling device, the ion trap and the ion cooling device each having an electrode assembly, the controller being configured to: Causing ions to be introduced into the ion trap from an upstream ion device that transfers ions within a selected range of m / z ratios, Causing ions having m / z ratios within a first range of m / z ratios to be introduced into the ion cooling device, Applying the first RF capture amplitude to the electrode array of the ion cooling device to capture introduced ions having m / z ratios within the first range of m / z ratios, Transferring trapped ions from the ion cooling device to the ion trap while applying the first RF trapping amplitude to the electrode assemblies of the ion cooling device and the ion trap, Applying the first RF trapping amplitude to the electrode array of the ion trap for a duration sufficient to allow cooling of the trapped ions; reducing the RF capture amplitude applied to the electrode array of the ion trap from the first RF capture amplitude to a second, lower RF capture amplitude; and either causing ions having m / z ratios within a second range of m / z ratios to be introduced and trapped in the ion cooling device by applying the second, lower RF trapping amplitude to the electrode assembly of the ion cooling device; or causing ions having m / z ratios within a second range of m / z ratios to be introduced and trapped in the ion trap by applying the second, lower RF trapping amplitude to the electrode assembly of the ion trap. [23] The controller of claim 22, wherein when the controller is configured to cause ions having m / z ratios within a second range of m / z ratios to be introduced and trapped in the ion cooling device by applying the second, lower RF trapping amplitude to the electrode assembly of the ion cooling device, the controller is also configured to transfer the ions having m / z ratios within the second range of m / z ratios from the ion cooling device to the ion trap while applying the second RF trapping amplitude to the ion cooling device and to the ion trap. [24] A controller for controlling the capture of ions in an ion trap assembly, the ion trap assembly comprising an electrode assembly, the controller being configured to: Causing ions to be introduced into the ion trap assembly; Applying a first RF trapping frequency to the electrode assembly to trap introduced ions having m / z ratios within a first range of m / z ratios for a duration sufficient to allow cooling of the trapped introduced ions; Increasing the RF trapping frequency applied to the electrode assembly from the first RF trapping frequency to a second, higher RF trapping frequency that traps introduced ions with m / z ratios within a second range of m / z ratios, wherein a lower mass limit of the second mass range is below the lower mass cut-off of the ion trap assembly when the first RF trapping frequency is applied, wherein the controller is configured to determine a threshold value for a total number of trapped ions within the ion trap assembly in dependence on the first and second RF trapping frequencies, wherein the controller is further configured to control the application of the first and second RF trapping frequencies such that a total number of trapped ions in the ion trap assembly is maintained below the threshold. [25] A controller for controlling the fragmentation and capture of ions in an ion trap assembly, the ion trap assembly comprising an electrode assembly and configured to fragment ions, the controller being configured to: Causing the introduction of precursor ions into the ion trap arrangement, Causing fragmentation of the introduced precursor ions to generate product ions; Applying a first RF capture amplitude to the electrode array to capture product ions having m / z ratios within a first range of m / z ratios for a duration sufficient to allow cooling of the captured product ions; Reducing the RF capture amplitude applied to the electrode array from the first RF capture amplitude to a second, lower RF capture amplitude that captures product ions with m / z ratios within a second range of m / z ratios, wherein a lower mass limit of the second range of m / z ratios is below the lower mass cut-off of the ion trap arrangement when the first RF trapping amplitude is applied, wherein the controller is configured to determine a threshold for the total number of trapped ions within the ion trap assembly as a function of the first and second RF trap amplitudes, wherein the controller is further configured to control the application of the first and second RF trap amplitudes such that a total number of trapped ions in the ion trap assembly is maintained below the threshold. [26] Ion trap assembly comprising: an electrode arrangement; and the control according to one of claims 19 to 25. [27] Mass spectrometer comprising: an ion source configured to generate ions; an ion trap assembly configured to receive the ions generated by the ion source; and the control according to one of claims 19 to 25.

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