Radiant energy – Ionic separation or analysis – Methods
Reexamination Certificate
2002-10-09
2004-04-06
Lee, John R. (Department: 2881)
Radiant energy
Ionic separation or analysis
Methods
C250S282000, C250S287000, C250S286000, C250S281000
Reexamination Certificate
active
06717131
ABSTRACT:
FIELD OF THE INVENTION
The invention relates to methods and instruments for measuring daughter-ion spectra in reflector time-of-flight mass spectrometers with post-acceleration of parent and daughter ions selected by means of a parent-ion selector.
BACKGROUND OF THE INVENTION
The mass-to-charge ratio m/z of ions can be determined from their flight time in a time-of-flight mass spectrometer. For the sake of simplicity, only the mass m and its determination will be referred to in the following even though the measurement of the mass to charge ratio m/z, where z is the number of elementary charges carried by the ion, is always used in mass spectrometry. Since many types of ionization, such as Matrix-Assisted Laser Desorption and Ionization (MALDI), in the main produce only singly charged ions (z=1), the differences between m/z and m cease to be relevant for these forms of ionization.
The daughter ion or fragment ion spectra of parent ions which are selected by an ion selector on the basis of their time of flight can be measured in a time-of-flight mass spectrometer which has been fitted with an ion selector and a velocity-focusing reflector. The decomposition of parent ions into daughter ions or fragment ions can be produced by two different processes: firstly, by introducing excess energy while they are being ionized during laser bombardment (‘LID’ Laser Induced Decomposition), where so-called ‘metastable’ ions are produced which partially decompose as they travel through the mass spectrometer and secondly, by Collisionally Induced Decomposition (CID), which essentially leads to spontaneous decomposition, where somewhat different fragmentation rules prevail, such as the loss of side chains.
The reflector which has now gained general acceptance is the Mamyrin velocity-focusing, two stage reflector. In the first deceleration stage of the reflector, the ions are decelerated sharply, but in the second stage only weakly. Faster ions with the same mass as slower ions penetrate further into the relatively weak linear deceleration field of the second stage and therefore cover a somewhat longer distance which, by correct adjustment of the two deceleration fields, can compensate for the faster speed of the ions with the same mass emerging from a primary focus so that they arrive at the secondary focus at exactly the same time.
As well as the velocity focusing of ions with the same mass, there is energy dispersion for ions with the same velocity but with different masses. If the parent ions and the daughter ions which arise from the decomposition of parent ions enter the reflector simultaneously and with the same velocities, and therefore with different mass-dependent energy levels, they will be dispersed in the reflector by their different energies in accordance with their masses. This dispersion can be used for measuring the daughter ions.
However, the method of detecting daughter ions or fragment ions using reflectors such as these has serious disadvantages. When focusing is reasonably good, only ions with a relatively low relative energy range can be detected—in the standard commercially available instruments, approximately 25-30% of the energy (and mass) range under adjustment. Thus, for a medium-sized peptide, approximately 10-15 spectral segments have to be acquired if the entire fragment spectrum of the low masses of individual ionized amino acids up to the mass of the parent ions is to be measured. All these spectral segments have to be co-ordinated with each other using a complex mass-calibration process. Only then can the spectral segments be assembled in a data system to produce an artificially generated composite spectrum.
In the patent DE 198 56 014 C2 (U.S. Pat. No. 6,300,627), ways are described for recording daughter-ion spectra in a time-of-flight mass spectrometer with a two-stage reflector using a single non-segmented spectral scan. The patent also discloses other information regarding the MALDI ionization method and velocity focusing by delayed acceleration in the ion source. This method not only saves time when scanning the spectrum but also requires lower sample consumption, and this means that the sensitivity is significantly higher.
The method described in patent DE 198 56 014 C2 is based on the fact that the selected ions are post-accelerated so that they all fit within the energy window of the reflector. One of the methods suggested consists of only slightly accelerating the ions in an ion source with delayed acceleration, allowing them to decompose in an initial drift region, very rapidly raising them to a second acceleration potential using a potential cell (‘potential lift’) and accelerating them in subsequent acceleration sections to a second drift region. The second drift region can be at the same potential as the first drift region. In the preferred embodiment, the two drift regions are operated at ground potential. In the second drift region, very light ions then have a minimum energy which provides the second acceleration potential; the parent ions which have not decomposed have a maximum energy corresponding to the sum of the first and second acceleration. If a reflector is able to reflect particles with energy deviations corresponding to approximately 30% of the maximum energy, and if about 70% of the total energy is provided by the second acceleration potential, then the reflector can reflect all the daughter ions simultaneously. In this case scanning of the entire daughter-ion spectrum without any segmentation is possible.
At the same time, the potential lift itself can be used as the selector for selecting the parent ions for the daughter-ion spectrum. For much better results, an additional selector must be used, which will provide significantly better time resolution for the parent ions.
A prior art, optimized method for scanning daughter-ion spectra using time-of-flight mass spectrometers will appear as follows.
Ions are produced by the formation of a vapor cloud by laser bombardment focused onto a sample carrier of a solid sample containing analyte molecules embedded in a matrix substance. Due to the explosive expansion of the cloud in the vacuum, the ions have slightly different location-dependent, initial velocities. After a time delay, an acceleration field is switched on where, according to patent U.S. Pat. No. 5,654,545, for example, temporal focusing of the ions with a single mass but different initial speeds can be achieved at an adjustable location. The intermediate velocity is focused exactly on the location of the parent-ion separator, where the parent ions arrive, time-focused and at exactly the same time, so that high mass resolution can be achieved for the selection of a parent ion type. According to patent DE 196 38 577 C2 (U.S. Pat. No. 5,969,348), by using a temporal pulse profile for the acceleration field, it is possible to have the focal points for all the masses at the same location, i.e. in the parent-ion selector.
By means of a high energy density at the focal point of the laser, a high proportion of metastable ions ranging from a few tenths to a few percent can be achieved. In other words, these ions decompose to daughter ions and neutral particles as they travel through the mass spectrometer with a certain half-life. The daughter ions travel at the same speed as the parent ions. They are allowed through by the parent-ion separator at the same time as the parent ions but all other ions are masked out.
On leaving the parent-ion separator, the parent ions and daughter ions travel on to the potential lift and then drift slightly apart again due to small difference in the initial velocities. When they are in the potential lift cell, the potential is raised in a very short time. From here, they travel into the first acceleration path, which at this point has no field and is at the same potential. When transfer of the ions into the field-free acceleration path is complete, an acceleration potential is applied which, because of the location and velocity correlation of these ions, temporally focuses the ions of the same mass
Holle Armin
La Rotta Aurelio
Bruker Daltonik GmbH
Hashmi Zia R.
Lee John R.
LandOfFree
Clean daughter-ion spectra using time-of-flight mass... does not yet have a rating. At this time, there are no reviews or comments for this patent.
If you have personal experience with Clean daughter-ion spectra using time-of-flight mass..., we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Clean daughter-ion spectra using time-of-flight mass... will most certainly appreciate the feedback.
Profile ID: LFUS-PAI-O-3207657