After a precursor ion has been captured within an ion trap (2), electrons having a high energy equal to or higher than 30 eV are introduced from an electron irradiator (7) into the ion trap (2) to increase the number of charges of the ion through an interaction between the electrons and the ion. Hydrogen radicals are subsequently introduced from a hydrogen radical irradiator (5) into the ion trap (2) to dissociate the ion by a hydrogen-attachment dissociation (HAD) method. The larger the number of charges of the ion is, the higher the dissociation efficiency by the HAD method becomes. Therefore, for example, even in the case of using an ion source in which most of the generated ions are singly charged ions as in a MALDI ion source, the dissociation efficiency can be improved by increasing the number of charges of the precursor ion within the ion trap (2).

A dissociation device fragments a precursor ion, producing at least two different product ions with overlapping m/z values in the dissociation device. The dissociation device applies an AC voltage and a DC voltage creating a pseudopotential that traps ions below a threshold m/z including the at least two product ions. The dissociation device receives a charge reducing reagent that causes the trapped at least two product ions to be charge reduced until their m/z values increase above the threshold m/z set by the AC voltage. The increase in the m/z values of the at least two product ions decreases their overlap. The at least two product ions with increased m/z values are transmitted to another device for subsequent mass analysis by applying the DC voltage to the dissociation device relative to a DC voltage applied to the other device.

In one aspect, a method of performing mass spectrometry is disclosed, which comprises forming at least two adduct ions of a molecular species in a sample, wherein said adduct ions comprise at least two different isotopes (e.g., the naturally occurring isotopes) of any of sodium and potassium, subjecting the adduct ions to Electron Impact Excitation of Ions from Organics (EIEIO) fragmentation to generate a plurality of fragment product ions, and using a mass analyzer to generate a mass spectrum of said product ions. The mass spectrum of the product ions can be analyzed to identify the parent ion. By way of example, the analysis of the mass spectrum can involve utilizing a reference library that contains mass spectral information regarding electron induced ionization of a plurality of reference compounds.

A mass isolation device selects a precursor ion of a sample that has been digested using a protease. A first fragmentation device fragments the precursor ion using collision-induced dissociation (CID), and the resulting product ions are analyzed using a mass analyzer producing a CID spectrum. A list of theoretical candidate glycopeptide sequences is determined from CID spectrum. The mass isolation device again selects the precursor ion of the sample. A second fragmentation device fragments the precursor ion using electron-based dissociation (ExD), and the resulting product ions are analyzed using the mass analyzer producing a CID spectrum. For each sequence of the list, the sequence is computationally fragmented, producing theoretical fragments, mass-to-charge ratio (m/z) values are calculated for the theoretical fragments, and the sequence is scored using c and z fragment matching rules. The highest scoring sequence is identified as a peptide sequence of a glycopeptide of the sample.

A system and method is described for characterizing glycopeptides which includes a first quadrupole mass filter, a multipole rod set of an ion guide, a lens electrode, an ExD device and a mass analyzer. The multipole rod set is adapted to receive a radial radio frequency (RF) trapping voltage and a radial dipole direct current (DC) voltage The lens electrode is adapted to receive an axial trapping alternating current (AC) voltage and a DC voltage. The ExD device performs electron capture dissociation or electron transfer dissociation, the ExD device being positioned so that an entrance of the ExD device is disposed on the other side of the lens electrode opposite the multipole rod set. The mass analyzer is positioned at an exit of the ExD device for receiving ions from the ExD device.

A mass spectrometer is operated to simultaneously measure precursor and production data over a number of acquisitions. For each acquisition, the following steps are performed. Ion transfer optics inject ions from an ion beam into an ELIT causing the ions to oscillate axially between two electric fields produced by two the sets of reflectrons. The ELIT measures a time domain image current of the oscillating ions from ion injection to a total acquisition time, Tacq1, and fragments the oscillating ions at one or both turning points of the oscillating ions adding product ions to the oscillating ions. The fragmentation is performed at a delay time relative to the ion injection that is increased by a time increment in each subsequent acquisition making the fragmentation dependent on ion position. The measured time domain image current is stored as a row or column of a two-dimensional matrix.

The present invention concerns a method for sequencing oligosaccharides, which makes it possible to identify the primary sequence of an oligosaccharide of unknown structure, including its monosaccharide composition, the position (regiochemistry) and configuration (stereochemistry) of glycosidic bonds, the nature and position of functional modifications, and its branched structure, particularly including the identification of the reducing end.

This invention discloses a method for constructing a set of database of one or more saccharides, a logical procedure for automatic determination of sequential mass spectra, and a method, program and system for determination the structures of oligosaccharides and glycoconjugates by the set of database. In one aspect, the sequential mass spectra measured by the method, program or system of the invention maybe instructed according to the logical procedure automatically or manually determined. By comparing the sequential mass spectra to the set of database, the structure of the carbohydrate comprising linkage position, anomeric configuration, composed monosaccharide and branch location of the carbohydrate sample can be identified. In another aspect, the method, program may be used to control one or more mass spectrometer automatically or manually.

A method and apparatus for analyzing samples using mass spectrometry are disclosed. The apparatus includes a reaction device configured to dissociate sample ions into fragments by reacting the sample ions with a charged species (e.g., electrons) such as through ECD, EID, or EIEIO. The kinetic energy of the charged species is such that the fragments may be detected and produce spectra that allow for the determination of isomeric species in the sample and the location of double bonds and/or the orientation of those double bonds within the sample molecules. The fragments may include radical fragments and non-radical fragments. Spectra resulting from analysis of the fragments may allow for the determination of the oxygen-radical fragments resulting from the dissociation of the sample molecules as confirmation of the presence of those radical fragments.

Methods are described for using a combination of mass spectroscopic and proteomic approaches for identifying the specific pairing of heavy and light chains for an intact antibody, an antibody fragment, a mixture of intact antibodies, or a mixture of antibody fragments.