Method of ion abundance augmentation in a mass spectrometer

A method of improving the detection limits of a mass spectrometer by: generating sample ions from an ion source; storing the sample ions in a first ion storage device; ejecting the stored ions into an ion selection device; selecting and ejecting ions of a chosen mass to charge ratio out of the ion selection device; storing the ions ejected from the ion selection device in a second ion storage device without passing them back through the ion selection device; repeating the preceding steps so as to augment the ions of the said chosen mass to charge ratio stored in the second ion storage device; and transferring the augmented ions of the said chosen mass to charge ratio back to the first ion storage device for subsequent analysis.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a National Stage application under 35 U.S.C. §371 of PCT Application No. PCT/GB2007/001362, filed Apr. 13, 2007, entitled “METHOD OF ION ABUNDANCE AUGMENTATION IN A MASS SPECTROMETER”, which claims the priority benefit of GB Application No. 0607542.8, filed Apr. 13, 2006, entitled “MASS SPECTROMETER WITH ION STORAGE DEVICE”, which applications are incorporated herein by reference in their entireties.

FIELD OF THE INVENTION

The present invention relates to a mass spectrometer and a method of mass spectrometry, in particular for performing MSnexperiments.

BACKGROUND TO THE INVENTION

Tandem mass spectrometry is a well known technique by which trace analysis and structural elucidation of samples may be carried out. In a first step, parent ions are mass analysed/filtered to select ions of a mass to change ratio of interest, and in a second step these ions are fragmented by, for example, collision with a gas such as argon. The resultant fragment ions are then mass analysed usually by producing a mass spectrum.

Various arrangements for carrying out multiple stage mass analysis or MSnhave been proposed or are commercially available, such as the triple quadrupole mass spectrometer and the hybrid quadrupole/time-of-flight mass spectrometer. In the triple quadrupole, a first quadrupole Q1acts as a first stage of mass analysis by filtering out ions outside of a chosen mass-to-charge ratio range. A second quadrupole Q2is typically arranged as a quadrupole ion guide arranged in a gas collision cell. The fragment ions that result from the collisions in Q2are then mass analysed by the third quadrupole Q3downstream of Q2. In the hybrid arrangement, the second analysing quadrupole Q3may be replaced by a time-of-flight (TOF) mass spectrometer.

In each case, separate analysers are employed before and after the collision cell. In GB-A-2,400,724, various arrangements are described wherein a single mass filter/analyser is employed to carry out filtering and analysis in both directions. In particular, an ion detector is positioned upstream of the mass filter/analyser, and ions pass through the mass filter/analyser to be stored in a downstream ion trap. The ions are then ejected from the downstream trap back through the mass filter/analyser before being detected by the upstream ion detector. Various fragmentation procedures, still employing a single mass filter/analyser, are also described, which permit MS/MS experiments to be carried out.

Similar arrangements are also shown in WO-A-2004/001878 (Verentchikov et al). Ions are passed from a source to a TOF analyser, which acts as an ion selector, from where ions are ejected to a fragmentation cell. From here, they pass back through the TOF analyser and are detected. For MSn, the fragment ions can be recycled through the spectrometer. US-A-2004/0245455 (Reinhold) carries out a similar procedure for MSnbut employs a high sensitivity linear trap rather than a TOF analyser to carry out the ion selection. JP-A-2001-143654 relates to an ion trap, ejecting ions on a circular orbit for mass separation followed by detection.

The present invention seeks against this background to provide an improved method and apparatus for MSn.

SUMMARY OF THE INVENTION

According to a first aspect of the present invention there is provided a method of improving the detection limits of a mass spectrometer comprising: (a) generating sample ions from an ion source; (b) storing the sample ions in a first ion storage device; (c) ejecting the stored ions into an ion selection device; (d) selecting and ejecting ions of a chosen mass to charge ratio out of the ion selection device; (e) storing the ions ejected from the ion selection device in a second ion storage device without passing them back through the ion selection device; (f) repeating the preceding steps (a) to (e) so as to augment the ions of the said chosen mass to charge ratio stored in the second ion storage device; and (g) transferring the augmented ions of the said chosen mass to charge ratio back to the first ion storage device for subsequent analysis.

This cycle may be repeated, optionally, multiple times, so as to allow MSn.

The present invention thus employs a cyclical arrangement in which ions are trapped, optionally cooled, and ejected from an exit aperture. A subset of these ions are returned to the ion storage device. This cyclical arrangement provides a number of advantages over the art identified in the introduction above, which instead employs a “back and forth” procedure via the same aperture in the ion trap. Firstly, the number of devices required to store and inject ions into the ion selector is minimised (and in the preferred embodiment is just one). Modern storage and injection devices that permit very high mass resolution and dynamic range are expensive to produce and demanding to control so that the arrangement of the present invention represents a significant cost and control saving over the art. Secondly, by using the same (first) ion storage device to inject into, and receive ions back from, an external ion selection device, the number of MS stages is reduced. This in turn improves ion transport efficiency which depends upon the number of MS stages. Typically, ions ejected from an external ion selector will have very different characteristics to those of the ions ejected from the ion storage device. By loading ions into the ion storage device through a dedicated ion inlet port (a first ion transport aperture), particularly when arriving back at the ion storage device from an external fragmentation device, this process can be carried out in a well controlled manner. This minimises ion losses which in turn improves the ion transport efficiency of the apparatus.

This technique also allows the detection limit of the instrument to be improved, where the ions of the chosen mass to charge ratio are of low abundance in the sample. Once a sufficient quantity of these low abundance precursor ions have been built up in the second ion storage device, they can be injected back to the first ion storage device for capture there (bypassing the ion selection device) and subsequent MSnanalysis, for example. Although preferably the ions leave the first ion storage device through a first ion transport aperture and are received back into it via a second separate ion transport aperture, this is not essential in this aspect of the invention and ejection and capture through the same aperture are feasible.

Optionally, at the same time as the low abundance precursor ions are being moved to the second ion storage device to improve total population of these particular precursor ions, the ion selection device may continue to retain and further refine the selection of other desired precursor ions. When sufficiently narrowly selected, these precursor ions can be ejected from the ion selection device and fragmented in a fragmentation device to produce fragment ions. These fragment ions may then be transferred to the first ion storage device, and MSnof these fragment ions may then be carried out or they may likewise be stored in the second ion storage device so that subsequent cycles may further enrich the number of ions stored in this way to again increase the detection limit of the instrument for that particular fragment ion.

In a second aspect, the present invention may reside in a method of improving the detection limits of a mass spectrometer comprising: (a) generating sample ions from an ion source; (b) storing the sample ions in a first ion storage device; (c) ejecting the stored ions into an ion selection device; (d) selecting and ejecting ions of analytical interest out of the ion selection device; (e) fragmenting the ions ejected from the ion selection device in a fragmentation device; (f) storing fragment ions in a second ion storage device without passing them back through the ion selection device; (g) repeating the preceding steps (a) to (f) so as to augment the fragment ions stored in the second ion storage device, and (h) transferring the augmented fragment ions back to the first ion storage device for subsequent analysis.

As above, ion ejection from the first ion storage device and ion capture back there may be through separate ion transport apertures or through the same one.

Ions in the first ion storage device may be mass-analysed either in a separate mass analyser, such as an Orbitrap as described in the above-referenced U.S. Pat. No. 5,886,346, or may instead be injected back into the ion selection device for mass analysis there.

An ion source may be provided to supply a continuous or pulsed stream of sample ions to the ion storage device. In one preferred arrangement, the optional fragmentation device may be located between such an ion source and the ion storage device instead. In either case, complicated MSnexperiments may be carried out in parallel by allowing division of (and, optionally, separate analysis of) sub populations of ions, either directly from the ion source or deriving from previous cycles of MS. This in turn results in an increase in the duty cycle of the instrument and can likewise improve the detection limits of it as well.

Although preferred embodiments of the invention may employ any ion selection device, it is particularly suited to and beneficial in combination with an electrostatic trap (EST). In recent years, mass spectrometers including electrostatic traps (ESTs) have started to become commercially available. Relative to quadrupole mass analysers/filters, ESTs have a much higher mass accuracy (parts per million, potentially), and relative to quadrupole-orthogonal acceleration TOF instruments, they have a much superior duty cycle and dynamic range. Within the framework of this application, an EST is considered as a general class of ion optical devices wherein moving ions change their direction of movement at least along one direction multiple times in substantially electrostatic fields. If these multiple reflections are confined within a limited volume so that ion trajectories are winding over themselves, then the resultant EST is known as a “closed” type. Examples of this “closed” type of mass spectrometer may be found in U.S. Pat. No. 3,226,543, DE-A-04408489, and U.S. Pat. No. 5,886,346. Alternatively, ions could combine multiple changes in one direction with a shift along another direction so that the ion trajectories do not wind on themselves. Such ESTs are typically referred to as of the “open” type and examples may be found in GB-A-2,080,021, SU-A-1,716,922, SU-A-1,725,289, WO-A-2005/001878, and US-A-20050103992 FIG. 2.

Of the electrostatic traps, some, such as those described in U.S. Pat. No. 6,300,625, US-A-2005/0,103,992 and WO-A-2005/001878 are filled from an external ion source and eject ions to an external detector downstream of the EST. Others, such as the Orbitrap as described in U.S. Pat. No. 5,886,346, employ techniques such as image current detection to detect ions within the trap without ejection.

Electrostatic traps may be used for precise mass selection of externally injected ions (as described, for example, in U.S. Pat. Nos. 6,872,938 and 6,013,913). Here, precursor ions are selected by applying AC voltages in resonance with ion oscillations in the EST. Moreover, fragmentation within the EST is achieved through the introduction of a collision gas, laser pulses or otherwise, and subsequent excitation steps are necessary to achieve detection of the resultant fragments (in the case of the arrangements of U.S. Pat. Nos. 6,872,938 and 6,013,913, this is done through image current detection).

Electrostatic traps are not, however, without difficulties. For example, ESTs typically have demanding ion injection requirements. For example, our earlier patent applications number WO-A-02/078046 and WO05124821A2 describe the use of a linear trap (LT) to achieve the combination of criteria required to ensure that highly coherent packets are injected into an EST device. The need to produce very short time duration ion packets (each of which contains large numbers of ions) for such high performance, high mass resolution devices means that the direction of optimum ion extraction in such ion injection devices is typically different from the direction of efficient ion capture.

Secondly, advanced ESTs tend to have stringent vacuum requirements to avoid ion losses, whereas the ion traps and fragmentors to which they may interface are typically gas filled so that there is typically at least 5 orders of magnitude pressure differential between such devices and the EST. To avoid fragmentation during ion extraction, it is necessary to minimise the product of pressure by gas thickness (typically, to keep it below 10−3. . . 10−2mm*torr), while for efficient ion trapping this product needs to be maximised (typically, to exceed 0.2 . . . 0.5 mm*torr)

Where the ion selection device is an EST, therefore, in a preferred embodiment of the present invention, the use of an ion storage device with different ion inlet and exit ports permits the same ion storage device to provide ions in an appropriate manner for injection into the EST, but nevertheless to allow the stream or long pulses of ions coming back from the EST via the fragmentation device to be loaded back into that first ion storage device in a well controlled manner, through the second or in certain embodiments, the third ion transport aperture.

Any form of electrostatic trap may be used, if this is what constitutes the ion selection device. A particularly preferred arrangement involves an EST in which the ion beam cross-section remains limited due to the focusing effect of the electrodes of the EST, as this improves efficiency of the subsequent ion ejection from the EST. Either an open or a closed type EST could be used. Multiple reflections allow for increasing separation between ions of different mass-to-charge ratios, so that a specific mass-to-charge ratio of interest may, optionally, be selected, or simply a narrower range of mass-to-charge ratios than was injected into the ion selection device. Selection could be done by deflecting unwanted ions using electric pulses applied to dedicated electrodes, preferably located in the plane of time-of-flight focus of ion mirrors. In the case of closed EST, a multitude of deflection pulses might be required to provide progressively narrowing m/z ranges of selection.

It is possible to use the fragmentation device in two modes: in a first mode, precursor ions can be fragmented in the fragmentation device in the usual manner, and in a second mode, by controlling the ion energy, precursor ions can pass through the fragmentation device without fragmentation. This allows both MSnand ion abundance improvement, together or separately: once ions have been injected from the first ion storage device into the ion selection device, specific low abundance precursor ions can be ejected controllably from the ion selection device and be stored back in the first ion storage device, without having been fragmented in the fragmentation device. This may be achieved by passing these low abundance precursor ions through the fragmentation device at energies insufficient to cause fragmentation. Energy spread could be reduced for a given m/z by employing pulsed deceleration fields (e.g. formed in a gap between two flat electrodes with apertures). When ions enter a decelerating electric field on the way back from the mass selector to the first ion storage device, higher energy ions overtake lower energy ions and thus move to a greater depth in the deceleration field. After all the ions of this particular m/z enter the deceleration field, the field is switched off. Therefore ions with initially higher energy experience a higher drop in potential relatively to ground potential than the lower energy ions, thus making their energies equal. By matching the potential drop to the energy spread upon exit from the mass selector, a significant reduction of the energy spread may be achieved. Fragmentation of ions may thereby be avoided, or, alternatively, control over the fragmentation may be improved.

In accordance with a further aspect of the present invention, there is provided a mass spectrometer comprising an ion storage device arranged to store ions, an ion selection device and a fragmentation/storage device. The ion selection device is arranged to receive ions stored in the first ion storage device and ejected therefrom, and to select a subset of ions from those received. The second fragmentation/storage device is arranged to receive at least some of the ions selected by the ion selection device. The second fragmentation/storage device is then configured, in use, to direct ions received from the ion selection device, or their products, back to the first ion storage device without passing them back through the ion selection device.

The present invention may also be found in a method of mass spectrometry comprising the steps of, in a first cycle, storing sample ions in a first ion storage device, the first ion storage device having an exit aperture and a spatially separate ion transport aperture; ejecting the stored ions out of the exit aperture into a separate ion selection device; receiving at least some of the ions ejected from the first ion storage device, or their derivatives, back through the ion transport aperture of the first ion storage device; and storing the received ions in the first ion storage device.

In accordance with a yet further aspect of the present invention, there is provided a method of mass spectrometry comprising storing ions in a first ion storage device; ejecting ions from the first ion storage device to an ion selection device; selecting a subset of ions within the ion selection device; ejecting the ions from the ion selection device; capturing at least some of the selected ions in one of a fragmentation device or second ion storage device; and returning at least some of the ions captured in the said one of the fragmentation device or second ion storage device respectively, or their products, to the first ion storage device along a return ion path that bypasses the ion selection device.

In accordance with still another aspect of the present invention there is provided a method of mass spectrometry comprising accumulating ions in an ion trap, injecting the accumulated ions into an ion selection device, selecting and ejecting a subset of the ions in the ion selection device, and storing the ejected subset of the ions directly back in the ion trap without intermediate ion storage.

Other preferred embodiments and advantages of the present invention will become apparent from the following description of a preferred embodiment.

DETAILED DESCRIPTION OF PREFERRED EMBODIMENTS

Referring first toFIG. 1, a mass spectrometer10is shown in block diagram format. The mass spectrometer10comprises an ion source20for generating ions to be mass analysed. The ions from the ion source20are admitted into an ion trap30which may, for example, be a gas-filled RF multipole or a curved quadrupole as is described, for example, in WO-A-05124821. The ions are stored in the ion trap30, and collisional cooling of the ions may take place as is described for example in our co-pending application number GB0506287.2, the contents of which are incorporated herein by reference.

Ions stored in the ion trap30may then be pulse-ejected towards an ion selection device which is preferably an electrostatic trap40. Pulsed ejection produces narrow ion packets. These are captured in the electrostatic trap40and experience multiple reflections therein in a manner to be described in connection particularly withFIG. 3below. On each reflection, or after a certain number of reflections, unwanted ions are pulse-deflected out of the electrostatic trap40, for example to a detector75or to a fragmentation cell50. Preferably, the ion detector75is located close to the plane of time-of-flight focus of the ion mirrors, where the duration of the ion packets is at a minimum. Thus, only ions of analytical interest are left in the electrostatic trap40. Further reflections will continue to increase the separation between adjacent masses, so that further narrowing of the selection window may be achieved. Ultimately, all ions having a mass-to-charge ratio adjacent to the mass-to-charge ratio m/z of interest are eliminated.

After the selection process is completed, ions are transferred out of the electrostatic trap40into the fragmentation cell50which is external to the electrostatic trap40. Ions of analytical interest that remain in the electrostatic trap40at the end of the selection procedure are ejected with sufficient energy to allow them to fragment within the fragmentation cell50.

Following fragmentation in the fragmentation cell, ion fragments are transferred back into the ion trap30. Here they are stored, so that, in a further cycle, a next stage of MS may be carried out. In this manner, MS/MS or, indeed, MSnmay be achieved.

An alternative or additional feature of the arrangement ofFIG. 1is that ions ejected from the electrostatic trap (because they are outside the selection window) may be passed through the fragmentation cell50without fragmentation. Typically, this could be achieved by decelerating such ions at relatively low energies so that they do not have sufficient energy to fragment in the fragmentation cell. These unfragmented ions which are outside of the selection window of immediate interest in a given cycle can be transferred onwards from the collision cell50to a auxiliary ion storage device60. In subsequent cycles (for example, when further mass spectrometric analysis of the fragment ions as described above has been completed), the ions rejected from the electrostatic trap40in the first instance (because they are outside of the selection window of previous interest) can be transferred from the auxiliary ion storage device60to the ion trap30for separate analysis.

Moreover the auxiliary ion storage device60can be used to increase the number of ions of a particular mass to charge ratio, particularly when these ions have a relatively low abundance in the sample to be analysed. This is achieved by using the fragmentation device in non-fragmentation mode and setting the electrostatic trap to pass only ions of particular mass to charge ratio that is of interest but which is of limited abundance. These ions are stored in the auxiliary ion storage device60but are augmented by additional ions of that same chosen mass to charge ratio selected and ejected from the electrostatic trap40using similar criteria in subsequent cycles. Ions of multiple m/z ratios could be stored together as well, e.g. by using several ejections from the trap40with different m/z.

Of course, either the previously unwanted precursor ions, or the precursor ions that are of interest but which have a low abundance in the sample and thus first need to be increased in number, can be the subject of subsequent fragmentation for MSn. In that case, the auxiliary ion storage device60could first eject its contents into the fragmentation cell50, rather than transferring its contents directly back to the ion trap30.

Mass analysis of ions can take place at various locations and in various ways. For example, ions stored in the ion trap may be mass-analysed in the electrostatic trap40(more details of which are set out below in connection withFIG. 2). Additionally or alternatively, a separate mass analyser70may be provided in communication with the ion trap30.

Turning now toFIG. 2, a preferred embodiment of a mass spectrometer10is shown in more detail. The ion source20shown inFIG. 2is a pulsed laser source (preferably a matrix-assisted laser desorption ionization (MALDI) source in which ions are generated through irradiation from a pulsed laser source22). Nevertheless, a continuous ion source, such as an atmospheric pressure electrospray source, could equally be employed.

Between the ion trap30and the ion source20is a pre-trap24which may, for example, be a segmented RF-only gas-filled multipole. Once the pre-trap is filled, ions in it are transferred into the ion trap30, which in the preferred embodiment is a gas-filled RF-only linear quadrupole, via a lens arrangement26. The ions are stored in the ion trap30until the RF is switched off and a DC voltage is applied across the rods. This technique is set out in detail in our co-pending applications, published as GB-A-2,415,541 and WO-A-2005/124821, the details of which are incorporated herein in their entirety.

The applied voltage gradient accelerates ions through ion optics32which may, optionally, include a grid or electrode34arranged to sense charge. The charge-sensing grid34permits estimation of the number of ions. It is desirable to have an estimate of the number of ions since, if there are too many ions, the resulting mass shifts become difficult to compensate. Thus, if the ion number exceeds a predefined limit (as estimated using the grid34), all ions may be discarded and an accumulation of ions in the pre-trap24may be repeated, with a proportionally lowered number of pulses from the pulsed laser22, and/or a proportionally shorter duration of accumulation. Other techniques for controlling the number of trapped ions could be employed, such as are described in U.S. Pat. No. 5,572,022, for example.

After acceleration through the ion optics32the ions are focused into short packets between 10 and 100 ns long for each m/z and enter the mass selector40. Various forms of ion selection device may be employed, as will become apparent from the following. If the ion selection device is an electrostatic trap, for example, the specific details of that are not critical to the invention. For example, the electrostatic trap, if employed, may be open or closed, with two or more ion mirrors or electric sectors, and with or without orbiting. At present, a simple and preferred arrangement of an electrostatic trap embodying the ion selection device40is shown inFIG. 3. This simple arrangement comprises two electrostatic mirrors42,44and two modulators46,48that either keep ions on a recurring path or deflect them outside of this path. The mirrors may be formed of either a circular or a parallel plate. As the voltages on the mirrors are static, they may be sustained with very high accuracy, which is favourable for stability and mass accuracy within the electrostatic trap40.

The modulators46,48are typically a compact pair of openings with pulsed or static voltages applied across them, normally with guard plates on both sides to control fringing fields. Voltage pulses with rise and fall times of less than 10-100 ns (measured between 10% and 90% of peak) and amplitudes up to a few hundred volts are preferable for high-resolution selection of precursor ions. Preferably, both modulators46and48are located in the planes of time-of-flight focusing of the corresponding mirrors42,44which, in turn, may preferably but do not necessarily coincide with the centre of the electrostatic trap40. Typically, ions are detected through image current detection (which is in itself a well known technique and is not therefore described further).

Returning again toFIG. 2, after a sufficient number of reflections and voltage pulses within the electrostatic trap40, only a narrow mass range of interest is left in the electrostatic trap40, thus completing precursor ion selection. Selected ions in the EST40are then deflected on a path that is different from their input path and which leads to the fragmentation cell50, or alternatively the ions may pass to detector75. Preferably, this diversion to the fragmentation cell is performed through a deceleration lens80which is described in further detail in connection withFIGS. 9 to 13below. The ultimate energy of the collisions within the fragmentation cell50may be adjusted by appropriate biasing of the DC offset on the fragmentation cell50.

Preferably, the fragmentation cell50is a segmented RF-only multipole with axial DC field created along its segments. With appropriate gas density in the fragmentation cell (detailed below) and energy (which is typically between 30 and 50 V/kDa), ion fragments are transported through the cell towards the ion trap30again. Alternatively or concurrently, ions could be trapped within the fragmentation cell50and then be fragmented using other types of fragmentation such as electron transfer dissociation (ETD), electron capture dissociation (ECD), surface-induced dissociation (SID), photo-induced dissociation (PID), and so forth.

Once the ions have been stored in the ion trap30again, they are ready for onward transmission towards the electrostatic trap40for a further stage of MSn, or towards the electrostatic trap40for mass analysis there, or alternatively towards the mass analyser70which may be a time-of-flight (TOF) mass spectrometer or an RF ion trap or FT ICR or, as shown inFIG. 2, an Orbitrap mass spectrometer. Preferably, the mass analyser70has its own automatic gain control (AGC) facilities, to limit or regulate space charge. In the embodiment ofFIG. 2, this is carried out through an electrometer grid90on the entrance to the Orbitrap70.

An optional detector75may be placed on one of the exit paths from the electrostatic trap40. This may be used for a multitude of purposes. For example, the detector may be employed for accurate control of the number of ions during a pre-scan (that is, automatic gain control), with ions arriving directly from the ion trap30. Additionally or alternatively, those ions outside of the mass window of interest (in other words, unwanted ions from the ion source, at least in that cycle of the mass analysis) may be detected using the detector. As a further alternative, the selected mass range in the electrostatic40may be detected with high resolution, following multiple reflections in the EST as described above. Still a further modification may involve the detection of heavy singly-charged molecules, such as proteins, polymers and DNAs with appropriate post-acceleration stages. By way of example only, the detector may be an electron multiplier or a microchannel/microsphere plate which has single ion sensitivity and can be used for detection of weak signals. Alternatively, the detector may be a collector and can thus measure very strong signals (potentially more than 104ions in a peak). More than one detector could be employed, with modulators directing ion packets towards one or another according to spectral information obtained, for example, from the previous acquisition cycle.

FIG. 4illustrates an arrangement which is essentially similar to the arrangement ofFIG. 2though with some specific differences. As such, like reference numerals denote parts common to the arrangements ofFIGS. 2 and 4.

The arrangement ofFIG. 4again comprises an ion source20which supplies ions to a pre-trap which in the embodiment ofFIG. 4is a auxiliary ion storage device60. Downstream of that pre-trap/auxiliary ion storage device60is a ion trap30(which in the preferred embodiment is a curved trap) and a fragmentation cell50. In contrast to the arrangement ofFIG. 2, however, the arrangement ofFIG. 4locates the fragmentation cell between the ion trap30and the auxiliary ion storage device60, that is, on the “source” side of the ion trap, rather than between the ion trap and the electrostatic trap as it is located inFIG. 2.

In use, ions are built up in the ion trap30and then orthogonally ejected from it through ion optics32to an electrostatic trap40. A first modulator/deflector100downstream of the ion optics32directs the ions from the ion trap30into the EST40. Ions are reflected along the axis of the EST40and, following ion selection there, they are ejected back to the ion trap30. To assist with ion guiding in that process, an optional electric sector (such as a toroidal or cylindrical capacitor)110may be employed. A deceleration lens is located between the electric sector110and the return path into the ion trap30. Deceleration may involve pulsed electric fields as described above.

Due to the low pressure in the ion trap30, ions arriving back at that trap30fly through it and fragment in the fragmentation cell50which is located between that ion trap30and the auxiliary ion storage device60(i.e. on the ion source side of the ion trap30). The fragments are then trapped in the ion trap30.

As withFIG. 2, an Orbitrap mass analyser70is employed to allow accurate mass analysis of ions ejected from the ion trap30at any chosen stage of MSn. The mass analyser70is located downstream of the ion trap (i.e. on the same side of the ion trap as the EST40) and a second deflector120“gates” ions either to the EST40via the first deflector100or into the mass analyser70.

Other components shown inFIG. 4are RF only transport multipoles that act as interfaces between the various stages of the arrangement as will be well understood by those skilled in the art. Between the ion trap30and the fragmentation cell50may also be located an ion deceleration arrangement (seeFIGS. 9-13below).

FIG. 5shows a further alternative arrangement to that shown inFIG. 2andFIG. 4and like components are once again labelled with like reference numerals. The arrangement ofFIG. 5is similar to that ofFIG. 2in that ions are generated by an ion source20and then pass through (or bypass) a pre-trap and auxiliary ion storage device60before being stored in a ion trap30. Ions are orthogonally ejected from the ion trap30, through ion optics32, and are deflected by a first modulator/deflector100onto the axis of an EST40, as withFIG. 4.

In contrast toFIG. 4, however, as an alternative to ion selection in the EST40, ions may instead be deflected by modulator/deflector100into an electric sector110and from there into a fragmentation cell50via an ion deceleration arrangement80. Thus (in contrast toFIG. 4) the fragmentation cell50is not on the source side of the ion trap30. Following ejection from the fragmentation cell50, ions pass through a curved transport multipole130and then a linear RF only transport multipole140back into the ion trap30. An Orbitrap or other mass analyser70is again provided to permit accurate mass analysis at any stage of MSn.

FIG. 6shows still a further alternative arrangement which is essentially identical in concept to the arrangement ofFIG. 2, except that the EST40is not of the “closed” type trap illustrated inFIG. 3, but is instead of the open type as is described in the documents set out in the introduction above.

More specifically, the mass spectrometer ofFIG. 6comprises an ion source20which provides a supply of ions to a pre-trap/auxiliary ion store60(further ion optics is also shown but is not labelled inFIG. 6). Downstream of the pre-trap/auxiliary ion storage device60is a further ion storage device which in the arrangement ofFIG. 6is once again a curved ion trap30. Ions are ejected from the curved trap30in an orthogonal direction, through ion optics32, towards an EST40′ where the ions undergo multiple reflections. A modulator/deflector100′ is located towards the “exit” of the EST40′ and this permits ions to be deflected either into a detector150or to a fragmentation cell50via an electric sector110and an ion decelerator arrangement80. From here, ions may be injected back into the ion trap30once more, again through an entrance aperture which is distinct from the exit aperture through which ions pass on their way to the EST40′. The arrangement ofFIG. 6also includes associated ion optics but this is not shown for the sake of clarity in that Figure.

In one alternative, the EST40′ ofFIG. 6may employ parallel mirrors (see, for example, WO-A-2005/001878) or elongate electric sectors (see, for example, US-A-2005/0103992). More complex shapes of trajectories or EST ion optics could be used.

FIG. 7shows still a further embodiment of a mass spectrometer in accordance with aspects of the present invention. As withFIG. 4, the spectrometer comprises an ion source20which supplies ions to a pre-trap which, as in the embodiment ofFIG. 4, is a auxiliary ion storage device60. Downstream of that pre-trap/auxiliary ion storage device60is a ion trap30(which in the preferred embodiment is a curved trap) and a fragmentation cell50. The fragmentation cell50could be located on either side of the ion trap30though in the embodiment ofFIG. 7the fragmentation cell50is shown between the ion source20and the ion trap30. As with the previous embodiments, an ion deceleration arrangement80is located in preference between the ion trap30and the fragmentation cell50.

In use, ions enter the ion trap30via an ion entrance aperture28and are accumulated in the ion trap30. They are then orthogonally ejected through an exit aperture29which is separate from the entrance aperture28, to an electrostatic trap40. In the arrangement shown inFIG. 7, the exit aperture is elongate in a direction generally perpendicular to the direction of ion ejection (i.e., the exit aperture29is slot-like). The ion position within the trap30is controlled so that the ions exit through one side (the left hand side as shown inFIG. 7) of the exit aperture29. Control of the position of the ions within the ion trap may be achieved in a number of ways, such as by applying differing voltages to electrodes (not shown) on the ends of the ion trap30. In one particular embodiment, ions may be ejected in a compact cylindrical distribution from the middle of the ion trap30whilst being recaptured as a much longer cylindrical distribution (as a result of divergence and aberrations within the system) of a much greater angular size.

Modified ion optics32′ are sited downstream of the exit from the ion trap30, and, downstream of that, a first modulator/deflector100″ directs the ions into the EST40. Ions are reflected along the axis of the EST40. As an alternative to the directing of the ions from the ion trap30into the EST40, the ions may instead be deflected by a deflector100″ downstream of the ion optics32′ into an Orbitrap mass analyser70or the like.

In the embodiment ofFIG. 7, the ion trap30operates both as a decelerator and as an ion selector. The extraction (dc) potential across the ion trap30is switched off and the trapping (rf) potential is switched on at the exact point at which ions of interest come to rest in the ion trap30following their return from the EST40. To inject into and eject from the EST40, the voltages on the mirror within the EST40(FIG. 3) which is closest to the lenses is switched off in a pulsed manner. After ions of interest are captured in the ion trap30, they are accelerated towards the fragmentation cell50on either side of the ion trap30, where fragment ions are generated and then trapped. After that, the fragment ions can be transferred to the ion trap30once more.

By ejecting ions from a first side of an elongate slot and capturing them back at or towards a second side of such a slot, the path of ejection from the ion trap30is not parallel to the path of recapture into that trap30. This in turn may allow injection of the ions into the EST40at an angle relative to the longitudinal axis of that EST40, as is shown in the embodiments ofFIGS. 4 and 5.

Of course, although a single slot-like exit aperture29is shown inFIG. 7, with ions exiting it towards a first side of that slot but being received back from the EST40via the other side of that slot, two (or more) separate but generally adjacent transport apertures (which may or may not then be elongate in the direction orthogonal to the direction of travel of ions through them) could instead be employed, with ions exiting via a first one of these transport apertures but returning into the ion trap30via an adjacent transport aperture.

Indeed, not only could the slot like exit aperture29ofFIG. 7be subdivided into separate transport apertures spaced in an generally orthogonal direction to the direction of travel of the ions during ejection and injection, but the curved ion trap30ofFIG. 7could itself be subdivided into separate segments. Such an arrangement is shown inFIG. 8.

The arrangement ofFIG. 8is very similar to that ofFIG. 7, in that the spectrometer comprises an ion source20which supplies ions to a pre-trap which is a auxiliary ion storage device60. Downstream of that pre-trap/auxiliary ion storage device60is a ion trap30′ (to be described further below) and a fragmentation cell50. As with the arrangement ofFIG. 7, the fragmentation cell50inFIG. 8could be located on either side of the ion trap30′ though in the embodiment ofFIG. 8the fragmentation cell50is shown between the ion source20and the ion trap30′, the ion trap30′ and the fragmentation cell50being separated by an optional ion deceleration arrangement80.

Downstream of the ion trap30is a first modulator/deflector100′″ which directs the ions into the EST40from an off axis direction. Ions are reflected along the axis of the EST40. To eject the ions from the EST40back to the ion trap30, a second modulator/deflector100″ in the EST40is employed. As an alternative to the directing of the ions from the ion trap30into the EST40, the ions may instead be deflected by the deflector100′″ into an Orbitrap mass analyser70or the like.

The curved ion trap30′ comprises in the embodiment ofFIG. 8, three adjoining segments36,37,38. The first and third segments36,38each have an ion transport aperture so that ions are ejected from the ion trap30′ via the first transport aperture in the first segment36, into the EST40, but are received back into the ion trap30′ via a second, spatially separate transport aperture in the third segment38. To achieve this, the same RF voltage may be applied to each segment of the ion trap30′ (so that in that sense the ion trap30′ acts as a single trap despite the several trap sections36,37,38) but with different DC offsets applied to each section so that the ions are not distributed centrally in the axial direction of the curved ion trap30′. In use, ions are stored in the ion trap30′. By suitable adjustment of the DC voltage applied to the ion trap segments36,37,38, ions are caused to leave the ion trap30′ via the first segment36for off axis injection into the EST40. The ions return to the ion trap30′ and enter via the aperture in the third segment38.

By maintaining the DC voltage on first and second segments36and37at a lower amplitude than the DC voltage applied to the third segment38when the ions are re-trapped from the EST40, the ions can be accelerated (eg by 30-50 ev/kDa) along the curved axis of the ion trap30′ so that they undergo fragmentation. In this manner the ion trap30′ is operable both as a trap and as a fragmentation device.

The resultant fragment ions are then cooled and squeezed into the first segment36by increasing the DC offset voltage on the second and third segments37,38relative to the voltage on the first segment36.

For optimal operation, fragmentation devices in particular require that the spread of energies of the ions injected into them is well controlled and held within a range of about 10-20 eV, since higher energies result in only low-mass fragments whereas lower energies provide little fragmentation. Many existing mass spectrometer arrangements, as well as the novel arrangements described in the embodiments ofFIGS. 1 to 7here, on the other hand, result in an energy spread of ions arriving at a fragmentation cell far in excess of that desirable narrow range. For example, in the arrangement ofFIGS. 1 to 7, the ions may spread in energy in the ion trap30,30′ due to spatial spread in that trap; due to space charge effects (e.g. Coulomb expansion during multiple reflections) in the EST40, and due to the accumulated effect of aberrations in the system.

In consequence some form of energy compensation is desirable.FIGS. 9 to 11show some specific but schematic examples of parts of an ion deceleration arrangement80for achieving that goal, andFIGS. 12 and 13show energy spread reduction and spatial spread for a variety of different parameters applied to such ion deceleration arrangements.

In order to achieve a suitable level of energy compensation, employing some of the embodiments described above, it is desirable to increase the ion energy dispersion. In other words, the beam thickness for a hypothetical monoenergetic ion beam is preferably smaller than the separation of two such hypothetical monoenergetic ion beams by the desired energy difference of 10-20 eV as explained above. Although a degree of energy dispersion could of course be achieved by physically separating the fragmentation cell50from the ion trap30or EST40by a significant distance (so that the ions can disperse in time), such an arrangement is not preferred as it increases the overall size of the mass spectrometer, requires additional pumping, and so forth.

Instead it is preferable to include a specific arrangement to allow deliberate energy dispersion without unduly increasing the distance between the fragmentation cell50and the component of the mass spectrometer upstream from it (ion trap30or EST40).FIG. 9shows one suitable device. InFIG. 9, an ion mirror arrangement200forming an optional part of the highly schematically represented ion deceleration arrangement80ofFIGS. 2-7is shown. The ion mirror arrangement200comprises an array of electrodes210terminating in a flat mirror electrode220. Ions are injected into the ion mirror arrangement from the EST40and are reflected by the flat mirror electrode220resulting in increased dispersion of the ions by the time they exit back out of the ion mirror arrangement and arrive at the fragmentation cell50. An alternative approach to the introduction of energy dispersion is shown inFIG. 11and described further below.

Once the degree of energy dispersion has been increased for example with the ion mirror arrangement200ofFIG. 9, ions are next decelerated. In general terms this may be achieved by applying a pulsed DC voltage to a decelerating electrode arrangement such as that illustrated inFIG. 10and labelled250. The decelerating electrode arrangement250ofFIG. 10comprises an array of electrodes with an entrance electrode260and an exit electrode270between which is sandwiched a ground electrode280. Preferably the entrance and exit electrodes are combined with differential pumping sections so as to reduce the pressure gradually between the (upstream) ion mirror arrangement200at a relatively low pressure, the decelerating electrode arrangement250at an intermediate pressure, and the relatively higher pressure required by the (downstream) fragmentation cell50. By way of example only, the ion mirror arrangement200may be at a pressure of around 10−8mBar, the decelerating electrode arrangement250may have a lower pressure limit of around 10−5mBar rising to around 10−4mBar via differential pumping, with a pressure in the range of 10−3to 10−2mBar or so in the fragmentation cell50. To provide pumping between the exit of the decelerating electrode arrangement250and the fragmentation cell50, an additional RF only multipole such as, most preferably, an octapole RF device, could be employed. This is shown inFIG. 11to be described below.

To achieve deceleration, DC voltages on one or both of the lenses260,270are switched. The time at which this occurs depends upon the specific mass to charge ratio of ions of interest. In particular, when ions enter a decelerating electric field, higher energy ions overtake lower energy ions and thus move to a greater depth in the deceleration field. After all the ions of this particular m/z enter the deceleration field, the field is switched off. Therefore ions with initially higher energy experience a higher drop in potential relatively to ground potential than the lower energy ions, thus making their energies equal. By matching the potential drop to the energy spread upon exit from the mass selector, a significant reduction of the energy spread may be achieved.

It will be understood that this technique permits energy compensation for ions of a certain range of mass to charge ratios, and not for an indefinitely wide range of different mass to charge ratios. This is because in a finite decelerating lens arrangement, only ions of a certain range of mass to charge ratios will be caused to undergo an amount of deceleration that can be matched to their energy spread. Any ions of widely differing mass to charge ratios to that selected will of course either be outside of the decelerating lens when it is switched, or likewise undergo a degree of deceleration but, having a largely different mass to charge ratio, the amount of deceleration will not then be balanced by the initial energy spread, i.e. the deceleration and penetration distance of higher energy ions will not then be matched to the deceleration and penetration distance of lower energy ions. Having said that, however, the skilled person will readily understand that this does not prohibit the introduction of ions of widely differing mass to charge ratios into the ion deceleration arrangement80, only that only ions of one particular range of mass to charge ratios of interest will undergo the appropriate degree of energy compensation to prepare them properly for the fragmentation cell50. Thus, the ions can either be filtered upstream of the ion deceleration arrangement80(so that only ions of a single mass to charge ratio of interest enter it in a given cycle of the mass spectrometer) or alternatively a mass filter can be employed downstream of the ion deceleration arrangement80. Indeed, it is even possible to use the fragmentation cell50itself to discard ions not of the mass to charge ratio of interest and which have been suitably energy compensated.

FIG. 11shows an alternative arrangement for decelerating ions and also optionally defocusing them as well. Here, the defocusing is achieved within the EST40(only a part of which is shown inFIG. 11) by pulsing the DC voltage on one of the electrostatic mirrors42,44(FIG. 3) at a time when ions of a mass to charge ratio of interest are in the vicinity of that electrostatic mirror42,44(because of the manner in which the EST40operates, the time at which ions of a particular m/z arrive at the electrostatic mirrors42,44is known). Applying a suitable pulse to that electrostatic mirror42or44results in that mirror42,44having a defocusing rather than a focusing effect on those ions.

Once defocused, the ions can then be ejected out of the EST by applying a suitable deflecting field to the deflector100/100′/100″. The defocused ions then travel towards a decelerating electrode arrangement300which decelerates ions of the selected m/z as explained above in connection withFIG. 10, by matching the initial energy spread to the drop in potential across the electric field defined by the decelerating electrode arrangement300.

Finally, ions exit the decelerating electrode arrangement300through termination electrodes310and pass through an exit aperture320into an octapole RF only device330to provide the desirable pumping described above.

FIGS. 12 and 13show plots of energy spread and spatial spread of ions of a specific mass to charge ratio, respectively, as a function of switching time of the DC voltage applied to the ion decelerating electrodes.

It can be seen fromFIG. 12that the reduction in energy spread achieved by an embodiment of the present invention can be as much as a factor of 20, reducing a beam with +/−50 eV spread to one of +/−2.4 eV. A longer switching time produces a smaller spatial spot size but a larger final energy spread with the particular decelerator system described here. The example is given here to show that beam characteristics other than energy spread must be considered, not to suggest that deceleration for optimal final energy spread always produces an increase in spatial spread of the final beam.

Other designs of decelerating lens used with other energy defocused beams could produce a still greater reduction in energy spread. Those skilled in the art will realise that there are many potential uses for the invention as a result. The use for which the invention was particularly addressed was that of improving the yield and type of fragment ions produced in a fragmentation process. As was noted earlier, for efficient fragmentation of parent ions, 10-20 eV ion energies are required, and clearly a great many ions in a beam having +/−50 eV energy spreads will be well outside that range. Ions having too high an energy predominantly fragment to low mass fragments which can make identification of the parent ion difficult, whilst a higher proportion of ions of low energy do not fragment at all. Without energy compensation, a parent ion beam having +/−50 eV energy spread directed towards a fragmentation cell would either produce a high abundance of low mass fragments, if all the beam were allowed to enter the fragmentation cell, or if only ions having the highest 20 eV of energy were allowed to enter (by use of a potential barrier prior to entry, for example) a great many ions would have been lost, and the process would be highly inefficient. The inefficiency would depend upon the energy distribution of the ions in the beam, with perhaps 90% of the beam being lost or unable to fragment due to insufficient ion energy.

By using the foregoing techniques, fragmentation of ions in the fragmentation cell may thereby be avoided if it is desired to pass ions through the fragmentation cell50(or store them there) in a given cycle of the mass spectrometer intact. Alternatively, control over the fragmentation may be improved when it is desired to carry out MS/MS or MS^n experiments.

Other uses for the ion deceleration technique described may be found in other ion processing techniques. Many ion optical devices can only function well with ions having energies within a limited energy range. Examples include electrostatic lenses, in which chromatic aberrations cause defocusing, RF multipoles or quadrupole mass filters in which the number of RF cycles experienced by the ions as they travel the finite length of the device is a function of the ion energy, and magnetic optics which disperse in both mass and energy. Reflectors are typically designed to provide energy focusing so as to compensate for a range of ion beam energies, but higher order energy aberrations usually exist and an energy compensated beam such as is provided by the present invention will reduce the defocusing effect of those aberrations. Again, those skilled in the art will realise that these are only a selection of possible uses for the described technique.

Returning now to the arrangements of FIGS.2and4-8, in general terms, effective operation of each of the gas-filled units shown in these Figures depends upon the optimum choice of collision conditions and is characterised by collision thickness P·D, where P is the gas pressure and D is the gas thickness traversed by ions (typically, D is the length of the unit). Nitrogen, helium or argon are examples of collision gases. In the presently preferred embodiment, it is desirable that the following conditions are approximately achieved:

In the pre-trap24, it is desirable that P·D>0.05 mm·torr, but is preferably <0.2 mm·torr. Multiple passes may be used to trap ions, as described in our co-pending Patent Application No. GB0506287.2.

The ion trap30preferably has a P·D range of between 0.02 and 0.1 mm·torr, and this device could also extensively use multiple passes.

For any auxiliary ion storage device60employed, the collision thickness P·D is preferably between 0.02 and 0.2 mm torr. On the contrary, it is desirable that the electrostatic trap40is sustained at high vacuum, preferably at or better than 10−8torr.

The typical analysis times in the arrangement ofFIG. 2are as follows:Storage in the pre-trap24: typically 1-100 ms; Transfer into the curved trap30: typically 3-10 ms;Analysis in the EST40: typically 1-10 ms, in order to provide selection mass resolution in excess of 10,000;Fragmentation in the fragmentation cell50, followed by ion transfer back into the curved trap30: typically 5-20 ms;Transfer through the fragmentation cell50into a second ion storage device60, if employed, without fragmentation: typically 5-10 ms; andAnalysis in a mass analyser70of the Orbitrap type: typically 50-2,000 ms.

Generally, the duration of a pulse for ions of the same m/z should be well below 1 ms, preferably below 10 microseconds, while a most preferable regime corresponds to ion pulses shorter than 0.5 microseconds (for m/z between about 400 and 2000). In alternative terms and for other m/z, the spatial length of the emitted pulse should be well below 10 m, and preferably below 50 mm, while a most preferable regime corresponds to ion pulses shorter than 5-10 mm. It is particularly desirable to employ pulses shorter than 5-10 mm when employing Orbitrap and multi-reflection TOF analysers.

Although one specific embodiment has been described, the skilled reader will readily appreciate that various modifications could be contemplated.