The present application is a non-provisional application of U.S. Provisional Patent Application No. 62/881,349 filed on Jul. 31, 2019, entitled “Method and Apparatus for Tandem Mass Spectrometry with MALDI-TOF Ion Source”. The entire contents of U.S. Provisional Patent Application No. 62/881,349 are herein incorporated by reference. This patent application is also related to U.S. Pat. No. 9,543,138 entitled “Ion Optical System for MALDI-TOF Mass Spectrometer” and to U.S. Pat. No. 8,735,810 entitled “Time-of-Flight Mass Spectrometer with Ion Source and Ion Detector Electrically Connected”. U.S. Pat. Nos. 9,543,138 and 8,735,810 are incorporated herein by reference.
The section headings used herein are for organizational purposes only and should not to be construed as limiting the subject matter described in the present application in any way.
The first practical time-of-flight (TOF) mass spectrometer (MS) was described by Wiley and McClaren more than 50 years ago. TOF mass spectrometers were generally considered to be only a tool for exotic studies of ion properties for many years. See, for example, “Time-of-Flight Mass Spectrometry: Instrumentation and Applications in Biological Research,” Cotter R J., American Chemical Society, Washington, D.C. 1997, for review of the history, development, and applications of TOF-MS in biological research.
Early TOF mass spectrometer systems included ion sources with electron ionization in the gas phase where a beam of electrons is directed into the ion source. The ions produced have a distribution of initial positions and velocities that is determined by the intersection of the electron beam with the neutral molecules present in the ion source. The initial position of the ions and their velocities are independent variables that affect the flight time of the ions in a TOF-MS. Wiley and McLaren developed and demonstrated methods for minimizing the contribution of each of these distributions. Techniques for minimizing the contribution of initial position are called “space focusing” techniques. Techniques for minimizing the contribution of initial velocity are called “time lag focusing” techniques. One important conclusion made by Wiley and McLaren is that it is impossible to simultaneously achieve both space focusing and velocity focusing. Optimization of these TOF mass spectrometers required finding the optimum compromise between the space focusing and velocity focusing distributions.
Many mass spectrometer applications require an accurate determination of the molecular masses and relative intensities of metabolites, peptides, and intact proteins in complex mixtures, which is challenging. Some known mass spectrometers utilize tandem mass spectrometry to provide information on the structure and sequence of many biological polymers and allow unknown samples to be accurately identified. Tandem mass spectrometers employ a first mass analyzer to produce, separate and select a precursor ion, and a second mass analyzer to fragment the selected ions and record the fragment mass spectrum from the selected precursor. As the applications for mass spectrometer instrumentation and associated data grow, new and improved tandem mass spectrometer methods and apparatus are needed.
The present teaching, in accordance with preferred and exemplary embodiments, together with further advantages thereof, is more particularly described in the following detailed description, taken in conjunction with the accompanying drawings. The skilled person in the art will understand that the drawings, described below, are for illustration purposes only. The drawings are not necessarily to scale, emphasis instead generally being placed upon illustrating principles of the teaching. The drawings are not intended to limit the scope of the Applicant's teaching in any way.
The present teaching will now be described in more detail with reference to exemplary embodiments thereof as shown in the accompanying drawings. While the present teachings are described in conjunction with various embodiments and examples, it is not intended that the present teachings be limited to such embodiments. On the contrary, the present teachings encompass various alternatives, modifications and equivalents, as will be appreciated by those of skill in the art. Those of ordinary skill in the art having access to the teaching herein will recognize additional implementations, modifications, and embodiments, as well as other fields of use, which are within the scope of the present disclosure as described herein.
Reference in the specification to “one embodiment” or “an embodiment” means that a particular feature, structure, or characteristic described in connection with the embodiment is included in at least one embodiment of the teaching. The appearances of the phrase “in one embodiment” in various places in the specification are not necessarily all referring to the same embodiment.
It should be understood that the individual steps of the methods of the present teachings can be performed in any order and/or simultaneously as long as the teaching remains operable. Furthermore, it should be understood that the apparatus and methods of the present teachings can include any number or all of the described embodiments as long as the teaching remains operable.
The advent of naturally pulsed ion sources such as CF plasma desorption ions source, static secondary ion mass spectrometry (SIMS), and matrix-assisted laser desorption/ionization (MALDI) ion sources has led to renewed interest in TOF mass spectrometers. Recent work in TOF mass spectrometry has focused on developing new and improved TOF instruments and software that take advantage of MALDI and electrospray (ESI) ionization sources. These ionization sources have removed the volatility barrier for mass spectrometry and have facilitated the use of mass spectrometers for many important biological applications.
It is desirable for mass spectrometers to provide an accurate determination of the molecular masses and relative intensities of metabolites, peptides, and intact proteins in complex mixtures. The use of tandem mass spectrometry provides information on the structure and sequence of many biological polymers. Tandem mass spectrometers employ a first mass analyzer to produce, separate and select a precursor ion, and a second mass analyzer to fragment the selected ions and record the fragment mass spectrum from the selected precursor. A wide variety of mass analyzers and combinations thereof for use in tandem mass spectrometry are known in the literature.
An important advantage of TOF-MS is that essentially all of the ions produced are detected, which is not the case for scanning MS instruments. In conventional MS-MS tandem instruments, all of the ions produced are not detected because each precursor is selected sequentially and all non-selected ions are lost. This limitation of conventional MS-MS tandem instruments can be overcome by selecting multiple precursors following each laser shot. Recording fragment spectra from each of the multiple selected precursors can partially overcome the loss of non-selected ions and dramatically improve speed and sample utilization without requiring the acquisition of raw spectra at a higher rate.
There are many types of tandem mass spectrometers known in the art. One particular type of tandem mass spectrometer is a Q-TOF tandem mass spectrometer.
In this Q-TOF tandem mass spectrometer 200, the laser beam strikes the ion source at an angle relative to the axis of the quadrupole, and passes between the poles of a quadrupole or hexapole ion guide. Ions and neutrals desorbed by the laser are transmitted to the ion guide with the ions being transmitted to the quadrupole analyzer and most of the neutral matrix molecules being deposited on the rods of the ion guide. There are at least two major problems with this configuration. Since the ion guides operate at relatively low voltage, a first problem is that deposition of matrix produces an insulating film that causes surface charging so that frequent cleaning of the ion guide is required to maintain acceptable performance. Secondly, essentially all of the ions produced by the laser, including those from the matrix, are transmitted into the quadrupole analyzer. Often the total intensity of matrix ions is 6 to 9 orders of magnitude greater than that of ions of interest. This can result in greatly reduced sensitivity due to the ‘chemical noise” generated by the matrix.
One feature of apparatus of the present teaching is that they overcome the known problems with Q-TOF tandem mass spectrometers 200. This advantage is achieved, at least in part, by replacing the MALDI source and quadrupole analyzer with a first mass spectrometer. Another feature of the apparatus of the present teaching is that it can be constructed to integrate with Q-TOF tandem mass spectrometer 200. Some embodiments of the apparatus of the present teaching can fit into the enclosure of the Q-TOF tandem mass spectrometer 200.
The first deflector 314 deflects a portion of the ions to a second deflector 316 that deflects the ions to an aperture 316. The system of two ion deflectors 314, 316 and aperture 318 are referred to as ion optics 320 and serves to separate the ions from neutrals and into a mass analyzer 322 that includes an ion detector 324 that detects the ions. This MALDI-TOF spectrometer instrument 300 has demonstrated performance that far exceeds that of any other MALDI-TOF currently available.
There is an ion decelerator 418 that takes in ions from the output of the ion optics 410 and decelerates them to an exit aperture 420. The beam deflectors 412, 414 are used to direct and adjust the ion beam position and direction for any mechanical misalignments such that they pass through the exit aperture 420 with maximum transmission. Said another way, the ion decelerator 418 transmits decelerated ions through an exit aperture 420 at an output of the decelerator. In some embodiments, the potential applied to the exit aperture 420 is the same as the potential of the sample plate 406. In some embodiments, that potential is a zero potential. The exit aperture 420 in some embodiments is less than 100 micrometers in diameter. The size of the ion beam at the exit aperture in some embodiments is controlled by a lens. In some embodiments, the lens is configured to minimize the ion beam diameter at the exit aperture 420.
A quadrupole mass filter 422 takes in ions that pass through the exit aperture 420. The quadrupole mass filter 422 acts as a timed ion selector. The timed ion selector isolates, or selects, ions over a narrow mass range. The selected ions at the output of the quadrupole mass filter 422 are sent to a fragmentation chamber 424 and then to a second mass analyzer 426 that performs a mass spectrometry analysis on the fragments generated in the fragmentation chamber 424. The fragmentation chamber 424 may be, for example, a quadrupole ion fragmentation chamber or an electron capture fragmentation chamber. In some embodiments, the fragmentation chamber 424 may include one or more ion traps. The second mass analyzer 426 may be, for example, a time-of-flight mass analyzer, an orthogonal time-of-flight mass analyzer, or a quadrupole mass analyzer. In some embodiments, the system 423 that includes the quadrupole mass filter 422, fragmentation chamber 424 and second mass analyzer 426 may be all or part of an existing mass spectrometer system that is integrated with the MALDI ion source 402 of the present teaching. In some embodiments, the quadrupole mass filter 422 is a separate system that integrated with the MALDI ion source 402 and a separate fragmentation chamber 424 and mass spectrometer 426.
The addition of the ion decelerator 418 and quadrupole mass filter 422, also referred to as an ion selector, overcomes all of the problems found with earlier MALDI Q-TOF instruments, such as the one shown in
In the embodiment of the tandem TOF mass spectrometer 400 of
The light from a laser 608 passes through a lens 610 that has a 75 mm focal length. A dimension 612 of the distance from the lens to the plane 614 where the sample resides is 75 mm, to match the focal length of the lens. A dimension 616 of the upper section in some embodiments is 125 mm. An X-Y alignment stage 618 is used to set the relative position of the ion optics 604 output and the decelerator 606 input. The two deflector electrode pairs 620, 622 separate the ions from the neutrals generated by the laser 608 pulses and direct them to the decelerator 606.
The pulse of ions travels through an evacuated field-free region 770 and enters a decelerating field 780 that is the mirror image of accelerating field 760. Decelerating field 780 arises from a voltage −V2 766 applied at a first decelerating electrode positioned at distance D 772 from the last accelerating electrode and a voltage −V1 762 applied at an electrode a distance D2 768 from the first decelerating electrode and a ground potential at an electrode positioned at a distance D1 764 from the previous electrode. In some embodiments, these potentials are applied, for example with two electrodes positioned in the ion decelerator 418 that is described in connection with
In some embodiments, the accelerating voltages V1 762 and V2 766 are applied continuously, and then ions produced from sample 720 will arrive at exit plate 794 with substantially the same velocity and position relative to the axis of the accelerator 408 that they possessed initially. In some embodiments, the diameter of aperture 792 is at least as large as the diameter of the laser pulse impinging on sample plate 720, and then the velocity and spatial distribution of the ions reaching aperture 792 is substantially the same as the initial velocity and spatial distribution. The flight time of ions from sample plate 720 to exit plate 794 depends on the mass-to-charge ratio and initial velocity of the ions according to equations well known in the art. If an accelerating potential is applied to aperture plate 796, then the ions may be transmitted to quadrupole mass filter 798 with energy higher than their initial energy.
Quadrupole mass filter 798 is tuned to transmit ions with predetermined mass-to-charge ratio. Selected ions 799 are transmitted through aperture plate 797 and enter, for example, the fragmentation chamber 424 (
Equivalents
While the Applicant's teaching is described in conjunction with various embodiments, it is not intended that the Applicant's teaching be limited to such embodiments. On the contrary, the Applicant's teaching encompass various alternatives, modifications, and equivalents, as will be appreciated by those of skill in the art, which may be made therein without departing from the spirit and scope of the teaching.
Number | Name | Date | Kind |
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8735810 | Vestal | May 2014 | B1 |
9543138 | Vestal et al. | Jan 2017 | B2 |
Number | Date | Country | |
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20210035792 A1 | Feb 2021 | US |
Number | Date | Country | |
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62881349 | Jul 2019 | US |