Mass spectrometer with high duty cycle

Abstract

A method of mass spectrometry comprising the steps of: a) providing a mass spectrometer having a time-of-flight (TOF) mass analyzer; b) performing a survey scan including isolating a packet of precursor ion species and mass analyzing the ions to obtain a first mass spectral data; c) determining a time window during which one of the precursor ion species, or fragment or product ions derived therefrom, was mass analyzed; and d) isolating another packet of precursor ion species and mass analyzing the ions to obtain a second mass spectral data, wherein a pusher of the TOF mass analyzer is pulsed according to a plurality of respective successive pulse sequences during a plurality of pulse sequence periods, each pulse sequence comprising a pulse sequence period. f) selecting from the second mass spectral data that was acquired during a time period corresponding to a time window of the survey scan to acquire selected data; g) repeating steps d) to f) at least one further time to obtain a plurality of sets of selected data; and combining the plurality of sets of selected data and decoding the combined data to acquire mass spectral data indicative of the mass-to-charge ratios of ions detected by the TOF mass analyzer.

Description

Technical Field

[0001] (Cross - reference to related applications) This application claims the priority and benefit of UK Patent Application No. 2204104.0 filed on March 23, 2022. The entire content of this application is incorporated herein by reference.

[0002] (Field of the Invention) The present invention generally relates to a mass spectrometer that repeatedly pulses ions into a separation region to separate the ions according to their mass - to - charge ratio.

Background Art

[0003] In both data - dependent acquisition (DDA) and tandem (MSMS) mass spectrometry, precursor ions are filtered so that only selected species of precursor ions pass downstream for analysis at any given time. Therefore, the mass spectrometer selects different species of precursor ions that pass downstream for analysis. This process continues until the desired species of precursor ions are analyzed. Generally, in order to analyze a sample as quickly as possible, it is desirable to sequentially select and transmit as many precursor ions per unit time as possible. However, in conventional devices such as quadrupole - time - of - flight (QToF) mass spectrometers, simply increasing the speed at which precursor species are selected results in a linear decrease in the sensitivity of the detected ions. For example, if the analyzer settings are changed such that the number of precursor species passing through increases from 25 different precursors per second to 50 different precursors per second, this results in the number of ions in each fragment ion scan being halved.

[0004] It is desirable to increase the overall duty cycle of such devices.

Summary of the Invention

[0005] A first aspect of this disclosure is a method of mass spectrometry, comprising the steps of: a) providing a time-of-flight (TOF) mass spectrometer having a mass spectrometer having an ion separator for separating ions according to their physicochemical properties, an ion detector, and a pusher that, when pulsed, pushes ions into the time-of-flight domain and delivers them to the detector; b) performing an investigation scan, wherein a separation cycle is performed in which packets of precursor ion species are separated in the ion separator such that precursor ion species having different values ​​of physicochemical properties elute from the ion separator at different times, and then mass spectrometry the separated precursor ion species, or fragment ions or product ions derived therefrom, in the TOF mass spectrometer to obtain first mass spectral data; c) determining from the first mass spectral data a time window in which one of the precursor ion species, or fragment ions or product ions derived therefrom, was mass spectrometryed by the TOF mass spectrometer; and d) so that precursor ion species having different values ​​of physicochemical properties elute from the ion separator at different times. Steps include: performing another separation cycle in which another packet of precursor ion species is separated in an ion separator; then e) mass spectrometry of the precursor ion species separated in step d), or fragments or product ions derived therefrom, in a TOF mass spectrometer to obtain second mass spectral data, wherein the mass spectrometry comprises pulsing a pusher according to a plurality of consecutive pulse sequences during a plurality of pulse sequence periods, each pulse sequence consisting of consecutive pushes arranged such that the duration between any pair of pushes in the pulse sequence is different from the duration between any other pair of pushes in the pulse sequence; f) selecting mass spectral data acquired during the period corresponding to the time window of the investigation scan from the second mass spectral data to obtain selected data; g) repeating steps d) to f) at least one more time so that a plurality of sets of the selected data are obtained; h) combining the plurality of sets of the selected data to obtain combined data; and then,i) A method is provided which includes the step of decoding the combined data to obtain mass spectral data representing the mass-to-charge ratio of ions detected by a TOF mass spectrometer.

[0006] As mass spectral data for the same precursor ion species (or fragment or product ions derived therefrom) are selected and combined from multiple separation cycles, the number of ions produced at a given time of flight (i.e., having a given mass-to-charge ratio) is more likely to reach the level required by the decoding process for determining the presence of a mass peak. This is in contrast to known techniques in which mass spectral data obtained from a single separation cycle are decoded. In such known techniques, ions may not be determined to be associated with a mass peak if they have low abundances. The novel technique disclosed herein avoids such drawbacks because it decodes the data only after it has been combined.

[0007] Step c) may include determining a first time delay between the start time of an isolation cycle in an investigation scan and the start time of the time window, but the method may also include setting the period in step f) to start after a second time delay from the start time of another isolation cycle in step d), where the first time delay is the same as the second time delay.

[0008] The time window may have the same duration as the period in step f).

[0009] The step of selecting mass spectral data in step f) may include selecting only a portion of the mass spectral data obtained in step e). Other unselected mass spectral data obtained in step e) may be discarded or not selected in order to be combined with other data.

[0010] The selected mass spectral data obtained in step f) may include mass spectral data obtained during several different pulse sequence periods in step e).

[0011] Multiple different pulse sequence periods may be consecutive (i.e., directly adjacent) pulse sequence periods.

[0012] The combined data may be decoded based on knowledge of the pulse sequences used in each of the multiple pulse sequence periods.

[0013] All of the multiple pulse sequences may consist of the same pulse sequence; that is, pulses of the same pattern may be used for each of the pulse sequences.

[0014] The decoding process may involve assigning data corresponding to detected ions to a mass-to-charge ratio, and determining that a mass-to-charge ratio peak has been detected only if a threshold number of ions are assigned to a given mass-to-charge ratio.

[0015] If fewer than a threshold number of ions are assigned to a given mass-to-charge ratio, the data (from the combined data) relating to those ions may be discarded.

[0016] Optionally, the step of determining the time window in step c) includes determining a time window in which only a single precursor ion species, or a fragment ion or product ion derived therefrom, reaches the TOF mass spectrometer and is mass-analyzed by the TOF mass spectrometer. Thus, the selected mass spectral data in step f) may consist only of the mass spectral data for that single precursor ion species, or for a fragment or product ion derived therefrom.

[0017] This method may include fragmenting or reacting a precursor ion species in a fragmentation or reaction device between steps d) and e), and the mass spectrometry in step e) may include mass spectrometry of the resulting fragment or product ion.

[0018] It should be understood that fragmenting or reacting any given precursor ion species can result in the formation of multiple different fragments or product ion species. A TOF mass spectrometer can simultaneously mass analyze all of the different fragments or product species derived from any given precursor species.

[0019] The analyzer can associate mass spectral data representing the mass-to-charge ratio of fragment or product ions with their precursor ions. This may be done by associating the fragment or product ions with precursor ions detected by a TOF mass spectrometer simultaneously with or with the same intensity profile as these fragment or product ions (since some precursor ions may remain even after the fragmentation or reaction step). Alternatively, if the precursor ions are mass filtered and different precursor ions are permeated into the fragmentation or reaction device at different times, the fragment or product ions may be associated with their precursor ions based on their detection time.

[0020] Therefore, the method may include the steps of separating precursor ions according to their physicochemical properties, fragmenting or reacting the precursor ions to produce fragments or product ions, passing the fragments or product ions through a mass spectrometer so that fragments or product ions originating from different precursor ions are passed through the mass spectrometer over different time windows, and performing mass spectrometry on the fragments or product ions.

[0021] The method may include the steps of: providing a mass filter for filtering ions between an ion separator and a fragmentation or reaction device, the mass filter for filtering ions so as to allow only ions having a limited value or range of values ​​for the mass-to-charge ratio to pass through at any given time; and controlling the mass filter so as ions elute from the ion separator, the value or range of values ​​changes, so as to allow different types of ions to pass through toward the fragmentation or reaction device at different times.

[0022] The mass filter may be controlled to vary the value or a range of the value over a cycle time synchronized with the separation cycle of step d).

[0023] The mass filter may be controlled in synchronization with the elution time of ions from the ion separator so that, at any given time, the mass filter is set to allow the expected mass-to-charge ratio to pass through the ion separator to the mass filter. For example, if the ion separator is an ion mobility separator, the passage time of a given mass-to-charge ratio ion through the ion separator can be estimated (since mobility and mass-to-charge ratio are associated) and used to control the mass filter so that that mass-to-charge ratio can be selectively allowed to pass through as it exits the ion separator.

[0024] Thus, the mass filter can be controlled to transmit one or more ion species, while other ion species that are desirably transmitted later for analysis are still moving through the ion separator towards the mass filter. Thus, relatively few ions are discarded by the mass filter and the sensitivity of the analysis is relatively high. However, separating ions in this way can result in relatively few ions reaching the TOF mass analyzer per unit time, which can lead to the problems described herein when using encoded frequent pulsing (EFP) techniques. The present disclosure overcomes this by combining EFP data in a new manner.

[0025] When ions are fragmented or reacted, the separation of precursor ions by the ion separator is preserved in the fragments or product ions, and fragments or product ions of different precursor ion species can reach the TOF mass analyzer over different respective time periods.

[0026] Each of the separation cycles described herein can be initiated by pulsing a packet of ions, which includes the precursor ion species, into the ion separator.

[0027] For example, ions can be accumulated in an ion accumulator and pulsed periodically into the ion separator. The ion separator can perform a separation cycle between each pulse, and the mass filter can be changed during each separation cycle, for example in synchronization with the ion separator, as described above.

[0028] The first physicochemical property may be ion mobility.

[0029] For example, the ion separator may be a drift time ion mobility separator having an electrode and a voltage source that maintains a static DC gradient that biases ions through or against a background gas so that the ions are separated according to their mobility through the gas. Alternatively, the ion separator may have an electrode and a voltage source configured to repeatedly move a DC or pseudo-potential barrier along the ion separator so as to bias ions through or against the gas within the ion separator so that the ions are separated according to their mobility through the gas.

[0030] Alternatively, the first physicochemical property may be, for example, the mass-to-charge ratio.

[0031] The step of mass analyzing during the interrogation scan of step b) may include pulsing the pusher in such a manner that the duration between any pair of adjacent pusher pulses is greater than the flight time from the pusher to the ion detector for the ions of the maximum mass-to-charge ratio pushed by the pusher. Thus, the data obtained by the TOF mass analyzer in a multi-push interrogation scan is not multiplexed and does not require decoding.

[0032] It will be understood that the interrogation scan may process ions in the same manner as the (non-interrogation scan) mode described to be performed following the interrogation scan, except that the TOF mass analyzer and TOF data are used differently in two modes. In other words, if the non-interrogation scan mode includes the steps of mass filtering and / or fragmentation / reaction described above, the interrogation scan mode can also include the corresponding steps. This ensures that the time window identified in the interrogation scan corresponds to the duration during which the one of the precursor ion species, or a fragment or product ion derived therefrom, is mass analyzed in the TOF mass analyzer during the non-interrogation scan.

[0033] A first aspect of this disclosure also provides a mass spectrometer comprising an ion separator for separating ions according to their physicochemical properties, an ion detector, a time-of-flight (TOF) mass spectrometer having a pusher for pushing ions into the time-of-flight domain and sending them to the detector when pulsed, and a control circuit configured to perform the method described herein.

[0034] This mass spectrometer may be configured to perform any of the methods described herein.

[0035] A second aspect of the present disclosure provides a method for mass spectrometry, the method comprising: providing a time-of-flight (TOF) mass spectrometer having an ion detector and a pusher that, when pulsed, pushes ions into the time-of-flight domain and delivers them to the detector; mass spectrometry of ions in the TOF mass spectrometer to obtain data relating to the ions, the mass spectrometry step comprising pulsing the pusher according to a pulse sequence consisting of consecutive pushes arranged such that the duration between any pair of pushes in the pulse sequence is different from the duration between any other pair of pushes in the pulse sequence; determining that one or more ion species arrive in the TOF mass spectrometer during only a portion of one or more pulse sequence periods of a pulse sequence period; combining only the data obtained by the mass spectrometer during such portions of one or more pulse sequence periods; and decoding the combined data.

[0036] It is recognized that if ion species reach the TOF mass spectrometer over a period shorter than the pulse sequence duration, it may be undesirable to combine all the TOF data acquired during that period before the decoding step. For example, combining TOF data acquired over the entire pulse sequence duration would result in combining more noise than necessary, since the ions were not present in the TOF mass spectrometer for the entire pulse sequence duration.

[0037] The method may include, before the step of mass spectrometry of ions, a step of performing a survey scan in which one or more ion species are mass-spectrometryed by a TOF mass spectrometer; a step of determining from the survey scan a time window in which one or more ion species reach the TOF mass spectrometer; and a step of using the time window to determine a portion of one or more pulse sequence periods of a pulse sequence period in which one or more ion species reach the TOF mass spectrometer.

[0038] A second aspect of the present disclosure also provides a TOF mass spectrometer comprising an ion detector, a pusher that, when pulsed, pushes ions into the time-of-flight domain and delivers them to the detector, and a control circuit, the control circuit configured to perform mass spectrometry on ions and acquire ion-related data by pulsing the pusher according to a plurality of consecutive pulse sequences during a plurality of pulse sequence periods, each pulse sequence consisting of consecutive pushes arranged such that the duration between any pair of pushes in the pulse sequence is different from the duration between any other pair of pushes in the pulse sequence, determining that one or more ion species arrive at the TOF mass spectrometer during only a portion of one or more pulse sequence periods of the pulse sequence period, combining only the data acquired by the mass spectrometer during that portion of one or more pulse sequence periods, and decoding the combined data to acquire mass spectral data representing the mass-to-charge ratio of one or more species.

[0039] The TOF mass spectrometer may be configured to perform any of the methods described in relation to a second aspect of this disclosure.

[0040] The disclosure also provides a mass spectrometer equipped with a TOF mass spectrometer, according to a second aspect of the disclosure.

[0041] A third aspect of the present invention provides a method for mass spectrometry, the method comprising: (i) providing a time-of-flight (TOF) mass spectrometer having an ion detector and a pusher that, when pulsed, pushes ions into the time-of-flight domain and delivers them to the detector; (ii) mass spectrometry of ions in the TOF mass spectrometer to obtain data relating to the ions, the mass spectrometry step comprising pulsing the pusher according to a plurality of consecutive pulse sequences during a plurality of pulse sequence periods, each pulse sequence consisting of consecutive pushes arranged such that the duration between any pair of pushes in the pulse sequence is different from the duration between any other pair of pushes in the pulse sequence; (iii) combining data acquired during a plurality of different pulse sequence periods to obtain combined data; and (iv) decoding the combined data to obtain mass spectral data representing the mass-to-charge ratio of the ions.

[0042] Since data from multiple pulse sequence periods are combined, the number of ions produced at a given time of flight (i.e., with a given mass-to-charge ratio) is more likely to reach the level required by the decoding process to determine the presence of a mass peak. This is in contrast to known techniques where data obtained from a single pulse sequence period is decoded. In such known techniques, ions may not be determined to be associated with a mass peak if they have low abundance. The novel technique disclosed herein avoids such drawbacks because it decodes the data only after it has been combined.

[0043] The combined data may be decoded based on knowledge of the pulse sequences used in each of the multiple pulse sequence periods.

[0044] All of the multiple pulse sequences may consist of the same pulse sequence; that is, pulses of the same pattern may be used for each of the pulse sequences.

[0045] This decoding process can assign data corresponding to detected ions to a mass-to-charge ratio, and determines that a mass-to-charge ratio peak has been detected only if a threshold number of ions have been assigned to a given mass-to-charge ratio.

[0046] If fewer than a threshold number of ions are assigned to a given mass-to-charge ratio, the data (from the combined data) relating to those ions may be discarded.

[0047] At least one ion species can reach the TOF mass spectrometer over a time window containing multiple different pulse sequence periods and be mass-analyzed by the TOF mass spectrometer.

[0048] Multiple different pulse sequence periods may be consecutive (i.e., directly adjacent) pulse sequence periods.

[0049] This method may combine data acquired during all (i.e., not just some) of the pulse sequence periods in which at least one ion species reaches the TOF mass spectrometer, and then perform a decoding step on the resulting combined data.

[0050] At least one ion species may reach the TOF mass spectrometer over a time window corresponding to at least one pulse sequence period, and data acquired by the mass spectrometer during at least one pulse sequence period may be combined with data acquired by the mass spectrometer during a first pulse sequence period that begins after a time has passed since the entry of the at least one ion species into the TOF mass spectrometer was stopped.

[0051] This method may include a step of fragmenting or reacting a precursor ion upstream of a TOF mass spectrometer, and the step of mass spectrometry of the ion in the TOF mass spectrometer may include a step of mass spectrometry of the resulting fragment or product ion.

[0052] It should be understood that fragmenting or reacting any given precursor species can result in the formation of multiple different fragments or product ion species. A TOF mass spectrometer can simultaneously analyze all of the different fragments or product species derived from any given precursor species.

[0053] The analyzer can associate mass spectral data representing the mass-to-charge ratio of fragment or product ions with their precursor ions. This may be done by associating the fragment or product ions with precursor ions detected by a TOF mass spectrometer simultaneously with or with the same intensity profile as these fragment or product ions (since some precursor ions may remain even after the fragmentation or reaction step). Alternatively, if the precursor ions are mass filtered and different precursor ions are permeated into the fragmentation or reaction device at different times, the fragment or product ions may be associated with their precursor ions based on their detection time.

[0054] This method may include the steps of separating ions according to their physicochemical properties, supplying the resulting separated ions to a mass spectrometer, and performing the mass spectrometry step on these ions.

[0055] This method may include the steps of: separating precursor ions according to their physicochemical properties; fragmenting or reacting the precursor ions to generate fragments or product ions; passing the fragments or product ions through a mass spectrometer so that fragments or product ions originating from different precursor ions are passed through the mass spectrometer over different time windows; and performing mass spectrometry on the fragments or product ions.

[0056] The physicochemical properties may be, for example, ion mobility or mass-to-charge ratio. For example, the separator may be a drift-time ion-mobility separator having electrodes and a voltage source that maintains a static DC gradient that biases ions through a background gas to separate them according to their mobility. Alternatively, the separator may have electrodes and a voltage source configured to repeatedly move a DC or pseudo-potential barrier along the ion separator to bias ions through or against the gas in the ion separator so that ions separate according to their mobility through the gas.

[0057] The method may include the steps of: providing a mass filter for filtering ions between a separator and a TOF mass spectrometer, which filters ions so as to allow only ions having a limited value or range of values ​​for the mass-to-charge ratio to pass through at any given time; and controlling the mass filter so as ions elute from the ion separator, to change the value or range of values ​​so as to allow different types of ions to pass through toward the mass spectrometer at different times.

[0058] The mass filter may be controlled in synchronization with the elution time of ions from the separator.

[0059] For example, a mass filter may be controlled in synchronization with the elution time of ions from the separator so that, at any given time, the mass filter is set to allow the expected mass-to-charge ratio to pass through the separator to the mass filter. For example, if the separator is an ion mobility separator, the passage time of a given mass-to-charge ratio ion through the separator can be estimated (since mobility and mass-to-charge ratio are associated) and used to control the mass filter so that that mass-to-charge ratio can be selectively allowed to pass through as it exits the separator.

[0060] Therefore, the mass filter can be controlled to allow one or more ion species to pass through, while other ion species that are desirable to pass through later for analysis still move towards the mass filter through the separator. Thus, relatively few ions are discarded by the mass filter, and the sensitivity of the analysis is relatively high. However, separating ions in this way can result in relatively few ions reaching the TOF mass spectrometer per unit time, which can lead to the problems described herein when using encoded frequent pulsed (EFP) techniques. This disclosure overcomes this by combining EFP data in a novel manner.

[0061] A mass filter may be placed between the separator and the fragmentation or reaction device that performs the fragmentation or reaction described above.

[0062] When ions are fragmented or reacted, the separation of precursor ions by a separator allows for preservation of the fragment or product ion, so that fragments or product ions of different precursor ions can reach the TOF mass spectrometer at different time points.

[0063] The method may include the step of pulsing packets of ions containing different types of ions into a separator in order to initiate the step of separating ions.

[0064] For example, ions may be accumulated in an ion accumulator and periodically pulsed in a separator. The separator performs a separation cycle between each pulse, and the mass filter may be changed during each separation cycle, for example, in synchronization with the separator, as described above.

[0065] The method may include, before the step of mass spectrometry of ions, a step of performing a survey scan in which one or more ion species are mass-spectrometryed by a TOF mass spectrometer; a step of determining from the survey scan a time window in which one or more ion species reach the TOF mass spectrometer; and a step of selecting a plurality of different pulse sequence periods based on the time window determined in the survey scan.

[0066] For example, at least a portion of each of several different pulse sequence periods may overlap with the time window. Alternatively, the method may include the steps of selecting at least one pulse sequence period in which one or more species reach the TOF mass spectrometer as one or more pulse sequence periods from several different pulse sequence periods, and selecting a first pulse sequence period that begins after a time when one or more species stop entering the TOF mass spectrometer as another pulse sequence period from said several different pulse sequence periods.

[0067] These techniques, which utilize survey scan data, ensure that data for the desired ion species is combined.

[0068] The method includes the steps of: mass-analyzing one or more additional ion species using a TOF mass spectrometer in an investigation scan; determining a further time window from the investigation scan for one or more additional ion species to reach the TOF mass spectrometer; determining from the further time window that the one or more additional ion species arrive at the TOF mass spectrometer during step (ii) of claim 1, specifically during a portion of one or more pulse sequence periods; combining only the data acquired by the mass spectrometer during such portion of one or more pulse sequence periods; and decoding the combined data.

[0069] It is recognized that if ion species reach the TOF mass spectrometer over a period shorter than the pulse sequence duration, it may be undesirable to combine all the TOF data acquired during that period before the decoding step. For example, combining TOF data acquired over the entire pulse sequence duration would result in combining more noise than necessary, since the ions were not present in the TOF mass spectrometer for the entire pulse sequence duration.

[0070] During the survey scan, the TOF mass spectrometer may operate in a mode such that the duration between any pair of adjacent pusher pulses is greater than or equal to the time of flight from the pusher to the detector of the largest mass-to-charge ratio ion pushed by the pusher.

[0071] Therefore, in multi-push survey scans, data obtained by the TOF mass spectrometer is not multiplexed and does not require decoding.

[0072] In embodiments where a survey scan is used, it will be understood that the survey scan can process ions in the same manner as the non-survey scan mode described above, except that the TOF mass spectrometer and TOF data are used differently in the two modes. In other words, if the non-survey scan separates and / or filters and / or fragments ions, the survey scan may also include a corresponding step. This ensures that the time at which ions are determined to reach the TOF mass spectrometer in the survey scan corresponds to the time at which ions reach the TOF mass spectrometer in the non-survey scan. This information can then be used to select which data to combine in the non-survey mode.

[0073] A third aspect of the present disclosure also provides a TOF mass spectrometer having an ion detector, a pusher that, when pulsed, pushes ions into the time-of-flight domain and delivers them to the detector, and a control circuit configured to perform mass spectrometry on ions and acquire data related to the ions by pulsing the pusher according to a plurality of consecutive pulse sequences during a plurality of pulse sequence periods, each pulse sequence consisting of consecutive pushes arranged such that the duration between any pair of pushes in the pulse sequence is different from the duration between any other pair of pushes in the pulse sequence, and is configured to combine the data acquired during a plurality of different pulse sequence periods to acquire combined data, and then decode the combined data to acquire mass spectral data representing the mass-to-charge ratio of the ions.

[0074] The TOF mass spectrometer may be configured to perform, for example, any of the methods described herein in relation to a third embodiment.

[0075] The Disclosure also provides a mass spectrometer comprising: an ion separator for separating ions according to their physicochemical properties; a mass filter for mass filtering the separated ions received from the ion separator, the mass filter being configured to allow ions with different mass-to-charge ratios to pass through as the ions elute from the ion separator; a fragmentation or reaction device for fragmenting or reacting the ions received from the mass filter to produce fragments or product ions; and the aforementioned TOF mass spectrometer for mass spectrometry of the fragments or product ions.

[0076] The mass spectrometer may be configured, for example, in relation to a third embodiment, to perform any of the methods described herein. [Brief explanation of the drawing]

[0077] [Figure 1]A schematic diagram of an embodiment of a mass spectrometer according to one embodiment of the present disclosure is shown. [Figure 2] This invention demonstrates how a mass filter can be controlled as a function of time to allow precursor ions to pass through, according to one embodiment of this disclosure. [Figure 3] The plot is the same as in Figure 2, except that the fragment or product ion species that can be generated from the precursor ion are also shown. [Figure 4] This example shows how ions can be mass-analyzed by a TOF mass spectrometer according to uncoded high-frequency pulsed technology. [Figure 5] This example shows how ions can be mass-analyzed by a TOF mass spectrometer according to coded high-frequency pulsed technology. [Figure 6] This invention illustrates a technique according to one embodiment of the present disclosure in which data acquired by a TOF mass spectrometer during multiple pulse sequence periods are combined before the data is decoded. [Figure 7] This invention illustrates a technique according to one embodiment of the present disclosure in which only data acquired by a TOF mass spectrometer during a portion of a pulse sequence period is combined before the data is decoded. [Modes for carrying out the invention]

[0078] Hereafter, various embodiments of the present invention will be described only as examples, with reference to the accompanying drawings. Figure 1 shows a schematic diagram of an embodiment of the mass spectrometer according to the present disclosure. The analyzer comprises an ion source 1, an ion accumulator 2, an ion separator 3, a mass filter 4, a fragmentation or reaction device 5, and a time-of-flight (TOF) mass spectrometer 6. It should be understood that other ion optics devices, such as one or more RF ion guides, electrostatic lenses, or impact-cooled RF ion guides, may also be provided within the analyzer, and are configured to bias ions through them by using an axial electric field or by moving a potential barrier along them.

[0079] During use, ions are supplied from ion source 1, which may be an ion source that generates ions from the analyte sample, or an ion optical device that supplies ions. Ions from ion source 1 may be accumulated in ion accumulator 2 and then released into separator 3 as ion packets. Accumulator 2 may accumulate ions and periodically pulse them into separator 3. Accumulator 2 may be configured to allow ions to enter accumulator 2 continuously, or it may prevent ions from entering accumulator 2 when ion packets are being pulsed into separator 3. In any case, accumulator 2 accumulates ions from ion source 1 in between the times when ion packets are being pulsed into separator 3. However, accumulator 2 may be omitted, and ion source 1 may instead pulse ions into separator 3, for example, by being a pulsed ion source, or by providing an ion gate between ion source 1 and separator 3.

[0080] The separator 3 is controlled such that each packet of ions it receives internally from the accumulator 2 is separated according to one or more physicochemical properties. For example, the separator 3 may separate ions according to ion mobility, mass-to-charge ratio, or a combination of both of these physicochemical properties. For example, the separator 3 may be an ion mobility separator that drives ions toward the gas so that they separate according to their mobility through the gas and dissolve from the separator in a time dependent on their mobility through the gas. The separator 3 may drive ions toward the gas using an electric field, such as one generated by a static DC gradient or one generated by moving a DC potential barrier along the device, in known ways. The gas may be substantially static, or the gas may flow in the opposite direction to the direction in which the ions are biased by the electric field.

[0081] Next, the separated ions pass from separator 3 to mass filter 4. Mass filter 4 is controlled to allow only a single mass-to-charge ratio (or a limited range of mass-to-charge ratios) to pass through at any given time, and as ions separated from the ion packet elute from separator 3, the single mass-to-charge ratio (or limited range) changes over time. Mass filter 4 may be modified to synchronize with the separation cycle of separator 3.

[0082] Figure 2 illustrates an example of how the mass filter 4 can be controlled as a function of time as ions separated from the ion packet elute from the separator 3. More specifically, Figure 2 shows a plot of the mass-to-charge ratio transmitted by the mass filter 4 as a function of time since the ion packet was pulsed into the separator 3. In this example, the mass filter 4 is controlled to transmit only ions with a first, relatively low mass-to-charge ratio at the first time. The mass filter is then controlled to transmit only ions with a second, higher mass-to-charge ratio at a second, later time. The mass filter is then controlled to transmit only ions with a third, higher mass-to-charge ratio at a third, later time. The mass filter is then controlled to transmit only ions with a fourth, lower mass-to-charge ratio at a fourth, later time. The mass filter is then controlled to transmit only ions with a fifth, higher mass-to-charge ratio at a fifth, later time.

[0083] The mass filter 4 may be, for example, a resolved quadrupole mass filter, but it is intended that other types of mass filters may be used. It will also be understood that the mass filter 4 may be controlled to transmit different mass-to-charge ratios at different times in ways other than those shown in the example in Figure 2.

[0084] After a delay period following the pulse of an ion packet into separator 3, another packet of ions may be pulsed into separator 3, and the above process may be repeated to separate and filter the ions in this next packet. The delay period may be the time it takes for all of the target ions in one ion packet (i.e., those that filter 4 is set to pass through at a certain point) to pass through separator 3 and reach filter 4, but may be set to a shorter or longer delay period if desired. Any desired number of ion packets may be separated sequentially and then filtered in the manner described above.

[0085] The separated ions that have been permeated by filter 4 (i.e., not removed by the filter) are transferred to a fragmentation or reaction device 5, where they may be fragmented or reacted to produce fragment or product ions. The fragmentation or reaction device 5 may be a collision-induced dissociation (CID) device, an electron capture dissociation (ECD) device, an electron transfer dissociation (ETD) device, a surface-induced dissociation (SID) device, or a device that fragments ions according to any other technique. For example, the device may react ions with reagent ions or molecules to fragment the ions, or the device may be used to react ions with reagent ions or molecules to form other types of product ions, such as adduct ions.

[0086] Figure 3 shows the same plot as Figure 2, except that the fragment or product ion species generated from the precursor ions are superimposed. In the illustrated example, the first precursor species, transmitted through mass filter 4, is shown as the first dark ellipse, and fragments or reacts to produce two types of fragments or product ions, which are shown by lighter colored ellipses above and below the first dark ellipse. In this example, one of the fragment or product ion species has a higher mass-to-charge ratio than the first precursor species, and the other has a lower mass-to-charge ratio. In this example, each of the second to fifth precursor species (represented by dark ellipses) fragments or reacts to produce three types of fragments or product ions (represented by light ellipses).

[0087] It should be understood that different precursor ions arrive at the fragmentation or reaction device 5 at different times due to their separation in separator 3, and therefore different precursor ions fragment or react at different times, producing their fragments or product ions at different times. Figure 3 shows that fragment or product ion species are produced over a time that precisely matches the time their precursor ion species are permeated through filter 4, however this may not be the case, and it should be understood that the time at which any given fragment or product ion species is produced may differ from the time that the precursor ion species was permeated through filter 4, due to, for example, the time it takes for the precursor ion species to permeate to (or through) the fragmentation or reaction device 5, and / or the time it takes for the precursor species to undergo fragmentation or reaction within device 5.

[0088] The separation of precursor ion species induced by separator 3 may be preserved among the fragments or product ion species derived from these precursor ion species. In particular, different precursor ion species arrive at the fragmentation or reaction device 5 at different times, and therefore, their respective fragments or product ion species are produced at corresponding different times. The fragments or product ion species may be biased out of the fragmentation or reaction device 5 so that fragments or product ion species derived from different precursor species arrive at the TOF mass spectrometer 6 at their respective different times.

[0089] Assuming lossless accumulation and lossless ion transfer in accumulator 2, the duty cycle and resulting sensitivity for both precursor ions, and therefore fragment or product ions, in the examples shown in Figures 2 and 3 are more than five times better than conventional MSMS methods that mass filter precursor ions without accumulating and separating ions upstream of the mass filter.

[0090] As described above, the fragments or product ions generated in the fragmentation or reaction device 5 are transferred to the TOF mass spectrometer 6. As is well known in the art, the TOF mass spectrometer 6 has a pusher that pulses ion packets into the time-of-flight domain (e.g., the field-free domain) and toward the ion detector. As the ions pass through the time-of-flight domain, they separate according to their mass-to-charge ratio and then collide with the ion detector. Thus, the separated ions reach the ion detector at different times, and the time it takes for an ion to reach the detector is related to its mass-to-charge ratio. The mass-to-charge ratio of a given ion can be determined from the duration between the time the ion was pulsed into the time-of-flight domain and the time it was detected by the ion detector. Thus, the TOF mass spectrometer 6 can acquire data from the signals detected by the detector, determine the mass-to-charge ratio and their intensities of the ions pulsed into the mass spectrometer 6, and form a mass spectrum.

[0091] Ions may be reflected by one or more ion mirrors between the pusher and the ion detector to provide a relatively long flight path through the time-of-flight domain. This allows ions with different mass-to-charge ratios to be separated to a greater extent as they travel through the time-of-flight domain, thus providing the TOF mass spectrometer 6 with higher mass resolution. Therefore, the TOF mass spectrometer 6 may be a multi-reflecting TOF (MRTOF) mass spectrometer that uses ion mirrors to reflect the ions multiple times as they drift from the pusher towards the ion detector.

[0092] Fragments or product ions derived from any given species of precursor ions reach the TOF mass spectrometer 6 within a limited period of time related to the time it takes for those precursor species to leave the separator 3.

[0093] Figure 4 shows an example of how fragment or product ions are mass-analyzed by a TOF mass spectrometer according to a conventional (uncoded high-frequency pulsing) approach. In this example, three fragment or product ions with three different mass-to-charge ratios are generated from a single precursor species in a fragmentation or reaction device 5 and then sent to a TOF mass spectrometer 6 over the same period of time. The three fragment or product ions reach the pusher of the TOF mass spectrometer 6 over a time window 8 shown in Figure 4. The TOF mass spectrometer 8 periodically pulses the pusher, and therefore the pusher repeatedly samples the fragment or product ions through the time window 8 and mass-analyzes them. The time during which the TOF pusher is pulsed is indicated by a vertical line 10 on the x-axis. These multiple pulses 10 of the TOF pusher during the duration 8 cause multiple TOF mass spectra to be generated for the fragment or product ions originating from each precursor ion. Figure 4 shows only fragment or product ions from one of the precursor species permeated by filter 4, but it should be understood that fragment or product ions from other precursor ion species permeated by filter 4 (at later or earlier times for each of the species) will also be mass-analyzed in the TOF mass spectrometer 6 in a corresponding manner (at later or earlier times for each of the species). This conventional data acquisition method has proven useful because, for example, it allows for accurate measurements in the separation time domain of separator 3, and thus ultimately enables the determination of physicochemical properties such as ion mobility and collision cross-section. This approach also allows processing software to group together fragment or product ions that are common to the same precursor species.

[0094] However, the approach shown in Figure 4 provides the TOF mass spectrometer 6 with a relatively low TOF sampling duty cycle. This is because a relatively long duration is provided between any given pusher pulse 10 and the following pusher pulse, which allows time for the ion with the heaviest mass-to-charge ratio pushed by the given pusher pulse to arrive at the ion detector of the TOF spectrometer 6, so that another packet of ions is pushed toward the ion detector by the next pusher pulse. This means that the duration between pusher pulses 10 is greater than or equal to the time of flight from the pusher to the detector for the largest mass-to-charge ratio to be detected. Thus, the sampling duty cycle decreases with respect to the square root of the largest mass-to-charge ratio to be detected. The TOF sampling duty cycle with such a technique is typically in the range of 0.1% to 50%. These values ​​are typical maximum duty cycles.

[0095] To increase the sampling duty cycle of the TOF mass spectrometer 6, it is known to use an encoded frequent pulsing (EFP) technique in which the pusher is operated such that the duration between adjacent pulses 10 of the pusher is less than the time of flight of the heaviest mass-to-charge ratio to be analyzed. In such a technique, the pusher is controlled to perform a sequence of consecutive pushes, but the duration between any pair of pushes in the sequence is temporally arranged such that the duration between any other pair of pushes in the sequence is different from the duration between any other pair of pushes in the sequence. The sequence of pushes may be repeated one or more times during mass spectrometry. Target ions pulsed into the TOF mass spectrometer 6 by different pushes arrive at the TOF detector in overlapping periods according to this EFP technique, but the TOF mass spectrometer can decode the resulting data using a decoding algorithm to determine which data is associated with each push by using the timing of the pushes in the sequence (since the duration between any pair of pushes in the sequence is unique). Thus, the TOF mass spectrometer 6 can convert the data into mass spectral data representing the mass spectrum of the detected ions.

[0096] Figure 5 shows the same example as shown in Figure 4, but with control of the pusher of the TOF mass spectrometer 6 using EFP technology. The time the pusher is pulsed is indicated by a vertical line 12 on the x-axis. The pusher is controlled to perform a pulse sequence of consecutive pushes during a pulse sequence period 14, all of which are temporally arranged such that the duration between any pair of pushes in the pulse sequence (i.e., within the pulse sequence period 14) is different from the duration between any other pair of pushes in the pulse sequence. The same pulse sequence may be repeated in consecutive pulse sequence periods 14. In Figure 5, the first push in each pulse sequence is highlighted by being shown as a thicker vertical line than other pushes in the pulse sequence. In this example, the pusher is controlled to perform a pulse sequence of eight consecutive pushes within each pulse sequence period 14. However, it will be understood that each pulse sequence may consist of more or fewer than eight pushes.

[0097] The initiating pushes of adjacent pulse sequences may be closer to or further apart than shown. For example, the first push of each pulse sequence in Figure 5 is selected to coincide with one of the pushes 10 shown in Figure 4, but this is for illustrative purposes only and is not required. However, preferably, the initiating pushes of adjacent pulse sequences are temporally separated by an amount corresponding to, or greater than, the time of flight from the pusher to the detector of the pusher with the largest mass-to-charge ratio that is desired to be mass-analyzed by a TOF mass spectrometer (e.g., pulsed by the pusher).

[0098] In the example shown in Figure 5, it can be seen that there are eight times more pushes than in the example shown in Figure 4. Therefore, this requires increasing the sampling duty cycle of the TOF mass spectrometer by eight times.

[0099] As described above, each sequence of push 12 is performed over a pulse sequence period 14. The data acquired by the TOF mass spectrometer 6 during this pulse sequence period 14 can later be decoded by a decoding algorithm to obtain mass spectral data representing the mass spectra of the ions detected by the detector.

[0100] While EFP technology helps increase the sampling duty cycle of the TOF mass spectrometer 6, it is recognized that problems can arise due to the way the decoding algorithm operates to obtain mass spectral data representing the mass spectrum. More specifically, the decoding algorithm defines a minimum threshold for the number of ions that must be detected in any given time of flight (from the pusher to the detector) before it is determined that a mass-to-charge ratio peak exists. Otherwise, the number of detected ions may be statistically meaningless. For example, the decoding algorithm may define the minimum threshold as 10 ions. Then, if the decoding algorithm determines that fewer than 10 ions were detected in any given time of flight within the pulse sequence time, the data for those ions is rejected and not assigned to the mass-to-charge ratio.

[0101] Embodiments of the present disclosure improve such techniques by combining (i.e., summing) data acquired by a TOF mass spectrometer 6 from multiple pulse sequence periods 14, and then applying a decoding algorithm to the combined data. The combined data may then be decoded using the decoding algorithm to generate mass spectral data representing the mass spectra of detected ions. Thus, ions representing mass peaks are less likely to be discarded, even if they are present within each pulse sequence period 14 at levels below a minimum threshold set by the decoding algorithm. This is because, when data from multiple pulse sequence periods 14 are combined, the number of ions occurring at a given time of flight (i.e., having a given mass-to-charge ratio) is more likely to increase to a level above the minimum threshold required by the decoding algorithm to determine the presence of mass peaks.

[0102] Figure 6 illustrates a technique according to one embodiment of the present disclosure. This technique is the same as that described in relation to Figure 5, except that data obtained by the TOF mass spectrometer 6 during multiple pulse sequence periods 14 are combined (i.e., summed) and then decoded to obtain mass spectral data representing the mass spectra of the detected ions. The data acquired during multiple pulse sequence periods 14 can be combined to produce a single set of summed mass spectral data, such as a single set of histogrammed data. In the example shown, data from five pulse sequence periods 14 are combined before decoding, but it will be understood that data from different numbers of pulse sequence periods may be combined.

[0103] Figure 6 shows that fragment or product ions pass through the TOF mass spectrometer 6 over a time window 8 spanning at least part of four consecutive pulse sequence periods 14, and that these ions no longer enter the TOF mass spectrometer 6 during the illustrated fifth pulse sequence period 14. However, since ions sampled by the push in the fourth pulse sequence period 14 reach the TOF ion detector during the fifth pulse sequence period 14, it may still be desirable to combine the data acquired during the fifth pulse sequence period 14 with the data acquired during all (or a subset) of the first through fourth pulse sequence periods 14. More generally, TOF data acquired during at least one or at least some of the pulse sequence periods 14 that occur when a first type of fragment or product ion is received by (and mass-analyzed by) the TOF mass spectrometer may be combined with TOF data acquired during the first pulse sequence period 14 that begins after the first type of fragment or product ion has stopped entering the TOF mass spectrometer 6. This combined data is then decoded.

[0104] When summing the TOF data acquired during pulse sequence periods 14, it may be desirable to exclude TOF data acquired during the last pulse sequence period 14 that ends before the first species of the fragment or product ion begins to enter the TOF mass spectrometer 6, before decoding it. This helps ensure that TOF data related to ions other than the first species of the fragment or product ion is not included in the combined data. Although the data relating to the first species of the fragment or product ion is described as being acquired and combined, it will be understood from the above description that multiple fragments or product ions may be generated from the same precursor species, and these multiple fragments or product ions may reach the TOF mass spectrometer 6 during the same time window 8. Therefore, data relating to all of these multiple species of fragments or product ions may be acquired and combined in the manner described above.

[0105] Figure 6 shows an example of how fragments or product ions derived from a first species of precursor ion can be mass-analyzed in the TOF mass spectrometer 6. It will be understood that the TOF mass spectrometer 6 can also mass-analyze fragments or product ions derived from other species of precursor ions received in the TOF mass spectrometer 6 (at an earlier or later time) in a corresponding manner. For example, fragments or product ions derived from a second, different precursor ion will reach the TOF mass spectrometer over a different, later time window (as shown, for example, in Figure 3). These fragments or product ions can then be analyzed in a manner corresponding to those described in relation to Figure 6.

[0106] To determine which pulse sequence period 14 should be used when summing TOF data for fragments or product ions derived from a particular precursor species, the methods disclosed herein can first identify the time window in which these fragments or product ions enter the TOF mass spectrometer 6. This may be done by performing a survey scan to determine the time window in which various different fragments or product ions reach the TOF mass spectrometer 6. For example, the survey scan may include an MS scan to determine which precursor ions are present in the analyte sample. The survey scan can then perform the method described in relation to Figures 1 to 4, with the filter 4 controlled to allow different precursor ions to pass through among the precursor ions detected at different times in the MS scan. Alternatively, the survey scan may perform the method described in relation to Figures 1 to 4 without first performing an MS scan to determine which precursor ions are present in the analyte sample. Furthermore, it is assumed that the ions do not need to be fragmented or reacted during the survey scan, but instead, the precursor ions may be detected by the TOF mass spectrometer for the reason that the time window in which the precursor ions are detected will correspond to the time window in which their fragments or product ions are detected.

[0107] During the investigation scan, the TOF mass spectrometer 6 may operate in non-EFP mode, i.e., the duration between consecutive pusher pulses is greater than or equal to the time of flight from the pusher to the detector for the largest mass-to-charge ratio desired to be mass-analyzed, as described, for example, in relation to Figure 4. The obtained TOF data can then be used to determine the time window in which fragments or product ions originating from each precursor ion transmitted through the filter 4 reach the TOF mass spectrometer 6.

[0108] Next, the method described in relation to Figures 1 to 3 can be performed again, and the obtained fragment or product ions can be mass-spected according to the EFP technique described in relation to Figure 6, i.e., the TOF data acquired during multiple pulse sequence periods 14 can be combined and then decoded. Since the time window in which each species of fragment or product ion reaches the TOF mass spectrometer 6 is known from the survey scan, the mass spectrometer 6 can use this information to determine the timing of the pulse sequence periods 14 that occur while any given precursor ion species fragment or product ion species reaches the TOF mass spectrometer 6. The mass spectrometer 6 then selects and combines the TOF data acquired during all or a subset of these pulse sequence periods 14. The mass spectrometer 6 may also combine this data with TOF data acquired during a first pulse sequence period 14 that begins after the time when these fragment or product ion species stopped entering the TOF mass spectrometer 6. The combined data is then decoded as described herein.

[0109] The technique described above is that the TOF data to be combined (before decoding) is obtained from product or fragment ions co-eluted from only a single precursor ion species permeated during only a single separation cycle of separator 3. In other words, the TOF data to be summed (before decoding) is the TOF data obtained during the elution of a single precursor species from a single separation cycle of separator 3. However, it is also intended that the TOF data to be combined (before decoding) may be TOF data obtained during the elution time of the same single precursor ion species during multiple separation cycles of separator 3. This TOF data obtained from multiple separation cycles can be combined and then decoded. This serves to combine a relatively large amount of spectral data before the combined data is decoded using the decoding algorithm, thereby overcoming the aforementioned problem using the minimum threshold set by the decoding algorithm.

[0110] According to these techniques, the investigation scan can be performed, for example, in the same manner as described above. The investigation scan may be used to determine the time window in which the target precursor species reaches the TOF mass spectrometer 6 with respect to the start time of the separator separation cycle (e.g., the time it takes for the ion packet to be pulsed into separator 3). Alternatively, the scan may be used to determine the time window in which the fragment / product ions of the target precursor ion species reach the TOF mass spectrometer 6 (e.g., with respect to the start time of the separator separation cycle).

[0111] Next, the method described in relation to Figures 1 to 3 can be performed, and the resulting fragment or product ions can be mass-analyzed according to the EFP technique described in relation to Figure 6. In other words, packets of precursor ions are separated in separator 3, filtered by mass filter 4, fragmented or reacted in fragmentation or reaction device 5, and then the resulting fragment or product ions are mass-analyzed by TOF mass spectrometer 6. Since the time window in which fragment or product ions derived from the target precursor ion species reach TOF mass spectrometer 6 is known from the survey scan, the mass spectrometer 6 can use this information to determine the timing of pulse sequence periods 14 that occur while these fragment or product ion species reach TOF mass spectrometer 6. The mass spectrometer 6 then selects TOF data obtained during all or a subset of these pulse sequence periods 14 to be combined with other data. As described above, the mass spectrometer 6 may also select TOF data acquired during a first pulse sequence period 14 that begins after the time when these fragment or product ion species stopped entering TOF mass spectrometer 6 to be added with other data.

[0112] Next, the above method is repeated by pulsed another packet of precursor ions into separator 3, separating the precursor ions in separator 3, filtering the ions eluting from separator 3 using mass filter 4, fragmenting or reacting the filtered ions in a reaction device 5, and then mass spectrometry of the resulting fragments or product ions in TOF mass spectrometer 6. The mass spectrometer 6 then again selects portions of TOF data corresponding to fragments or product ions derived from the target precursor ion species and combines them with previously selected TOF data.

[0113] The method described above may be repeated any number of times, and each time, TOF data corresponding to a fragment or product ion derived from the target precursor ion species is combined with previously selected TOF data. The combined data may then be decoded to obtain mass spectral data.

[0114] In the embodiments described above, each species of fragment or product ion is described as reaching the TOF mass spectrometer 6 over a time window longer than the pulse sequence period 14, for example, over a period longer than the time of flight from the pusher to the detector for the maximum mass-to-charge ratio that is desired to be mass-analyzed by the TOF mass spectrometer 6. However, some species of fragment or product ions may reach the TOF mass spectrometer 6 over a time window shorter than the pulse sequence period 14. This may occur, for example, because precursor species of such fragment or product species elute from the separator 3 over a relatively short duration.

[0115] Figure 7 shows an example in which three types of fragments or product ions (derived from the same precursor) reach the TOF mass spectrometer 6 over a time window 8 shorter than the pulse sequence period 14. As described above, the TOF mass spectrometer 6 operates according to EFP technology. The number of pushes 12 within each pulse sequence period 14 differs from that shown in Figure 6, but this is for illustrative purposes only, and as explained above, each pulse sequence period 14 may contain any desired number of pushes 12.

[0116] It is recognized that when ion species reach the TOF mass spectrometer 6 over a time window 8 shorter than the pulse sequence period 14, it is desirable not only not to combine TOF data acquired during multiple pulse sequence periods 14 before the decoding step, but also that it may be undesirable to combine all TOF data acquired during a single pulse sequence period 14 before the decoding step. For example, combining TOF data obtained over the entire pulse sequence period 14 would result in excessive noise data because ions were not present in the TOF mass spectrometer 6 throughout the entire pulse sequence period 14.

[0117] According to embodiments of the present disclosure, in such a situation, TOF data acquired during only a portion of a pulse sequence period 14 are combined before decoding. For example, TOF data acquired during a portion of the pulse sequence period 14 corresponding to at least a portion of the duration that the ion species enters the TOF mass spectrometer 6 may be combined before decoding. Less preferably, TOF data acquired during a pulse sequence period 14 after the ion species has stopped entering the TOF mass spectrometer 6 may be combined before the decoding step, because the ion species can still be received by the TOF detector for at least a portion of this time.

[0118] The duration for which an ion species enters the TOF mass spectrometer 6 can be determined from the survey scan in a manner corresponding to the survey scan described above. Thus, the mass spectrometer 6 can use this information to determine the pulse sequence period 14 within which the species reaches the TOF detector and the duration for which the data should be totaled.

[0119] While the present invention has been described with reference to preferred embodiments, it will be understood by those skilled in the art that various modifications in form and detail can be made without departing from the scope of the invention as described in the appended claims.

[0120] For example, although the mass filter 4 is described as stepping between different separate mass permeation windows as ions elute from the separator 3, it is intended that the mass filter 4 may instead have a progressively scanned mass permeation window. For example, the mass permeation window may increase by only one mass-to-charge ratio unit at a time. Alternatively, the mass filter 4 may have a mass permeation window configured to allow a range of mass-to-charge ratios (e.g., a range consisting of multiple Das) to pass through simultaneously. This mass permeation window may be progressively scanned, and each time the mass permeation window moves to allow a new mass range to pass through, the new mass range may still partially overlap with the previous mass range that the filter 4 was configured to allow to pass through.

[0121] Alternatively or additionally, although filter 4 is described as a mass-to-charge ratio filter, it is intended that the filter may filter ions according to another physicochemical property such as ion mobility or FAIMS.

[0122] Alternatively or additionally, although separator 3 is described as an ion-mobility separator, it is intended that the separator may separate ions according to other physicochemical properties such as the mass-to-charge ratio.

[0123] Furthermore, although the techniques described herein relate to summing and decoding TOF data associated with detected fragment or product ions, it is intended that the TOF data may instead relate to precursor ions (i.e., sample ions that have not been fragmented or reacted to form different ions). Alternatively, or further, it is intended that the techniques described herein may be performed without separating and / or filtering the ions.

Claims

1. A method of mass spectrometry, a) A step of providing a mass spectrometer having an ion separator for separating ions according to their physicochemical properties, an ion detector and a time-of-flight (TOF) mass spectrometer having a pusher, wherein the pusher, when pulsed, pushes ions into the time-of-flight domain and sends them to the ion detector, b) A step of performing an investigation scan, comprising: performing a separation cycle in which packets of precursor ion species are separated in the ion separator such that the precursor ion species having different physicochemical properties elute from the ion separator at different times; and then mass spectrometry the separated precursor ion species, or fragment ions or product ions derived therefrom, in the TOF mass spectrometer to obtain first mass spectral data; c) A step of determining a time window in which one of the precursor ion species, or a fragment or product ion derived therefrom, was mass-analyzed by the TOF mass spectrometer from the first mass spectral data, d) The step of performing another separation cycle in which another packet of precursor ion species is separated in the ion separator such that the precursor ion species having different values ​​of physicochemical properties elute from the ion separator at different times, and then e) A step of obtaining second mass spectral data by mass spectrometry of the precursor ion species separated in step d) or a fragment or product ion derived therefrom in the TOF mass spectrometer, wherein the mass spectrometry comprises pulsing the pusher according to a plurality of consecutive pulse sequences during a plurality of pulse sequence periods, each pulse sequence consisting of consecutive pushes arranged such that the duration between any pair of pushes in the pulse sequence is different from the duration between any other pair of pushes in the pulse sequence. f) A step of selecting mass spectral data acquired from the second mass spectral data during a period corresponding to the time window of the survey scan, and acquiring the selected data. g) The step of repeating steps d) to f) at least once more so that multiple sets of the selected data are obtained, h) The step of combining multiple sets of the selected data to obtain the combined data, and then i) The steps of decoding the combined data to obtain mass spectral data representing the mass-to-charge ratio of the ions detected by the TOF mass spectrometer, Methods that include...

2. The method according to claim 1, wherein step c) includes determining a first time delay between the start time of the isolation cycle in the survey scan and the start time of the time window, and the method includes setting the period in step f) to begin after a second time delay from the start time of another isolation cycle in step d), wherein the first time delay is the same as the second time delay.

3. The method according to claim 1, wherein the time window has the same duration as the period in step f).

4. The method according to claim 1, wherein the selected mass spectral data obtained in step f) includes mass spectral data obtained during a plurality of different pulse sequence periods in step e).

5. The method according to claim 1, wherein the combined data is decoded based on knowledge of the pulse sequences used in each of the plurality of pulse sequence periods.

6. The method according to claim 1, wherein the decoding assigns data corresponding to the detected ions to a mass-to-charge ratio, and determines that a mass-to-charge ratio peak has been detected only when a number of ions exceeding a threshold are assigned to a given mass-to-charge ratio.

7. The method according to claim 1, wherein between step d) and e) a precursor ion species is fragmented or reacted in a fragmentation or reaction device, and the mass spectrometry in step e) is performed by mass spectrometry of the resulting fragment or product ion.

8. The method according to claim 7, comprising the steps of: providing a mass filter for filtering ions between the ion separator and the fragmentation or reaction device, the mass filter for filtering ions so as to allow only ions having a limited value or range of values ​​for the mass-to-charge ratio to pass through at any given time; and controlling the mass filter so as ions elute from the ion separator to allow different types of ions to pass through toward the fragmentation or reaction device at different times, thereby changing the value or range of values.

9. The method according to claim 8, wherein the mass filter is controlled to vary the value or a range of the value over a cycle time synchronized with the separation cycle of step d).

10. The method according to claim 1, wherein the physicochemical property is ion mobility.

11. The method according to claim 1, wherein during the investigation scan of step b), the step of mass spectrometry includes pulsing the pusher in such a manner that the duration between any pair of adjacent pusher pulses is greater than or equal to the time of flight of the ion with the largest mass-to-charge ratio pushed by the pusher from the pusher to the ion detector.

12. A mass spectrometer, Ion separators for separating ions according to their physicochemical properties. A time-of-flight (TOF) mass spectrometer having an ion detector and a pusher that, when pulsed, pushes ions into the time-of-flight domain and sends them to the ion detector, and A control circuit configured to perform the method described in any one of claims 1 to 11, A mass spectrometer equipped with the following features.

13. A method of mass spectrometry, The step of providing a time-of-flight (TOF) mass spectrometer having an ion detector and a pusher that, when pulsed, pushes ions into the time-of-flight domain and sends them to the ion detector, A step of obtaining data relating to an ion by mass spectrometry in the TOF mass spectrometer, wherein the mass spectrometry involves pulsing the pusher according to a plurality of consecutive pulse sequences during a plurality of pulse sequence periods, each pulse sequence consisting of consecutive pushes arranged such that the duration between any pair of pushes in the pulse sequence is different from the duration between any other pair of pushes in the pulse sequence; The steps include determining that one or more ion species arrive at the TOF mass spectrometer during only a portion of each of one or more pulse sequence periods within the pulse sequence period, The steps include combining only the data acquired by the TOF mass spectrometer during each of the aforementioned parts of the one or more pulse sequence periods, The step of decrypting this combined data, Methods that include...

14. The step of performing a survey scan, in which one or more ion species are mass-analyzed by the TOF mass spectrometer, prior to the step of mass-analyzing the ions, The steps include determining the time window in which the one or more ion species reach the TOF mass spectrometer from the aforementioned survey scan, The steps of using the time window to determine the portion of one or more pulse sequence periods in which one or more ion species reach the TOF mass spectrometer during the pulse sequence period, The method according to claim 13, including the method described in claim 13.

15. TOF mass spectrometer, An ion detector, and a pusher that, when pulsed, pushes ions into the time-of-flight domain and sends them to the ion detector, A control circuit, By pulsing the pusher according to a plurality of consecutive pulse sequences during multiple pulse sequence periods, ions are mass-spectrometrically analyzed to obtain data related to the ions, and each pulse sequence consists of consecutive pushes arranged such that the duration between any pair of pushes in the pulse sequence is different from the duration between any other pair of pushes in the pulse sequence. It is determined that one or more ion species arrive at the TOF mass spectrometer during only a portion of each of one or more pulse sequence periods within the pulse sequence period. Combine only the data acquired by the TOF mass spectrometer during each of the aforementioned portion of the one or more pulse sequence periods. The combined data is decoded to obtain mass spectral data representing the mass-to-charge ratio of one or more ion species. A control circuit is configured as follows, A TOF mass spectrometer equipped with this feature.

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