Radiant energy – Ionic separation or analysis – Ion beam pulsing means with detector synchronizing means
Reexamination Certificate
1999-01-19
2002-02-19
Anderson, Bruce (Department: 2881)
Radiant energy
Ionic separation or analysis
Ion beam pulsing means with detector synchronizing means
C250S282000, C250S288000
Reexamination Certificate
active
06348688
ABSTRACT:
FIELD OF THE INVENTION
The invention relates generally to mass spectrometers and specifically to tandem mass spectrometers.
BACKGROUND OF THE INVENTION
Mass spectrometers vaporize and ionize a sample and determine the mass-to-charge ratio of the resulting ions. One form of mass spectrometer determines the mass-to-charge ratio of an ion by measuring the amount of time it takes a given ion to migrate from the ion source, the ionized and vaporized sample, to a detector, under the influence of electric fields. The time it takes for an ion to reach the detector, for electric fields of given strengths, is a direct function of its mass and an inverse function of its charge. This form of mass spectrometer is termed a time-of-flight mass spectrometer.
Recently time-of-flight (TOF) mass spectrometers have become widely accepted, particularly for the analysis of relatively nonvolatile biomolecules, and other applications requiring high speed, high sensitivity, and/or wide mass range. New ionization techniques such as matrix-assisted laser desorption/ionization (MALDI) and electrospray (ESI) have greatly extended the mass range of molecules which can be made to produce intact molecular ions in the gas phase, and TOF has unique advantages for these applications. The recent development of delayed extraction, for example, as described in U.S. Pat. Nos. 5,625,184 and 5,627,360, has made high resolution and precise mass measurement routinely available with MALDI-TOF, and orthogonal injection with pulsed extraction has provided similar performance enhancements for ESI-TOF.
These techniques provide excellent data on the molecular weight of samples, but little information on molecular structure. Traditionally tandem mass spectrometers (MS—MS) have been employed to provide structural information. In MS—MS instruments, a first mass analyzer is used to select a primary ion of interest, for example, a molecular ion of a particular sample, and that ion is caused to fragment by increasing its internal energy, for example, by causing the ion to collide with a neutral molecule. The spectrum of fragment ions is then analyzed by a second mass analyzer, and often the structure of the primary ion can be determined by interpreting the fragmentation pattern. In MALDI-TOF, the technique known as post-source decay (PSD) can be employed, but the fragmentation spectra are often weak and difficult to interpret. Adding a collision cell where the ions may undergo high energy collisions with neutral molecules enhances the production of low mass fragment ions and produces some additional fragmentation, but the spectra are difficult to interpret. In orthogonal ESI-TOF, fragmentation may be produced by causing energetic collisions to occur in the interface between the atmospheric pressure electrospray and the evacuated mass spectrometer, but currently there is no means for selecting a particular primary ion.
The most common form of tandem mass spectrometry is the triple quadrupole in which the primary ion is selected by the first quadrupole, and the fragment ion spectrum is analyzed by scanning the third quadrupole. The second quadrupole is typically maintained at a sufficiently high pressure and voltage that multiple low energy collisions occur. The resulting spectra are generally rather easy to interpret and techniques have been developed, for example, for determining the amino acid sequence of a peptide from such spectra. Recently hybrid instruments have been described in which the third quadrupole is replaced by a time-of-flight analyzer.
Several approaches to using time-of-flight techniques both for selection of a primary ion and for analysis and detection of fragment ions have been described previously. For example, a tandem instrument incorporating two linear time-of-flight mass analyzers using surface-induced dissociation (SID) has been used to produce the product ions. In a later version, an ion mirror was added to the second mass analyzer.
U.S. Pat. No. 5,206,508 discloses a tandem mass spectrometer system, using either linear or reflecting analyzers, which is capable of obtaining tandem mass spectra for each parent ion without requiring the separation of parent ions of differing mass from each other. U.S. Pat. No. 5,202,563 discloses a tandem time-of-flight mass spectrometer comprising a grounded vacuum housing, two reflecting-type mass analyzers coupled via a fragmentation chamber, and flight channels electrically floated with respect to the grounded vacuum housing. The application of these devices has generally been confined to relatively small molecules; none appears to provide the sensitivity and resolution required for biological applications, such as sequencing of peptides or oligonucleotides.
For peptide sequencing and structure determination by tandem mass spectrometry, both mass analyzers must have at least unit mass resolution and good ion transmission over the mass range of interest. Above molecular weight 1000, this requirement is best met by MS—MS systems consisting of two double-focusing magnetic deflection mass spectrometers having high mass range. While these instruments provide the highest mass range and mass accuracy, they are limited in sensitivity, compared to time-of-flight, and are not readily adaptable for use with modern ionization techniques such as MALDI and electrospray. These instruments are also very complex and expensive.
SUMMARY OF THE INVENTION
The invention relates to tandem time-of-flight mass spectrometry including: (1) an ion generator; (2) a timed ion selector in communication with the ion generator (3) an ion fragmentation chamber in communication with the ion selector; and (4) an analyzer in communication with the fragmentation chamber. In one embodiment, the ion generator comprises a pulsed ion source in which the ions are accelerated so that their velocities depend on their mass-to-charge ratio. The pulsed ion source may comprise a laser desorption ionization or a pulsed electrospray source. In another embodiment, the ion generator comprises a continuous ionization source such as a continuous electrospray, electron impact, inductively coupled plasma, or a chemical ionization source. In this embodiment, the ions are injected into a pulsed ion source in a direction substantially orthogonal to the direction of ion travel in the drift space. The ions are converted into a pulsed beam of ions and are accelerated toward the drift space by periodically applying a voltage pulse.
In one embodiment, the timed ion selector comprises a field-free drift space coupled to the pulsed ion generator at one end and coupled to a pulsed ion deflector at another end. The drift space may include a beam guide confining the ion beam near the center of the drift space to increase the ion transmission. The pulsed ion deflector allows only those ions within a selected mass-to-charge ratio range to be transmitted through the ion fragmentation chamber. In one embodiment, the analyzer is a time-of-flight mass spectrometer and the fragmentation chamber is a collision cell designed to cause fragmentation of ions and to delay extraction. In another embodiment, the analyzer includes an ion mirror.
A feature of the present invention is the use of the fragmentation chamber not only to produce fragment ions, but also to serve as a delayed extraction ion source for the analysis of the fragment ions by time-of-flight mass spectrometry. This allows high resolution time-of-flight mass spectra of fragment ions to be recorded over their entire mass range in a single acquisition. Another feature of the present invention is the addition of a grid which produces a field free region between the collision cell and the acceleration region. The field free region allows the ions excited by collisions in the collision cell time to complete fragmentation.
The invention also relates to the measurement of fragment mass spectra with high resolution, accuracy and sensitivity. In one embodiment, the method includes the steps of: (1) producing a pulsed source of ions; (2) selecting ions of a specific range of mass-to-charge rati
Anderson Bruce
Perseptive Biosystems
Testa Hurwitz & Thibeault LLP
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