US008507849B2

(12) United States Patent (10) Patent No.: US 8,507,849 B2 BrOWn (45) Date of Patent: Aug. 13, 2013

(54) (56) References Cited (75) Inventor: Jeffery Mark Brown, Cheshire (GB) U.S. PATENT DOCUMENTS 6,300,627 B1 10/2001 Koster et al. (73) Assignee: Micromass UK Limited, Manchester 6,512,226 B1 1/2003 Loboda et al. (GB) 6,534,764 B1 3/2003 Verentchikov et al. 6,627,877 B1 9, 2003 Davis et al. 6,900,430 B2 5/2005 Okumura et al. (*) Notice: Subject to any disclaimer, the term of this 2002/0027194 A1 3/2002 Holle et al...... 250,287 patent is extended or adjusted under 35 2004/0232327 A1 11/2004 Bateman et al. U.S.C. 154(b) by 804 days. 2005/0242279 A1* 11/2005 Verentchikov ...... 250,287 2007/0034794 A1 2/2007 Cotter et al...... 250,287 (21) Appl. No.: 11/721,755 * cited by examiner PCT Fled: Dec. 19, 2005 (22) FOREIGN PATENT DOCUMENTS (86) PCT NO.: PCT/GB2OOS/OO4911 EP 1306881 5, 2003 WO 2004O21386 8, 2003 S371 (c)(1), WO 20040O8481 1, 2004 (2), (4) Date: Jul. 3, 2009 OTHER PUBLICATIONS (87) PCT Pub. No.: WO2006/064280 Bateman, R. H. et al; A Combined Magnetic Sector-Time-of-flight Mass Spectrometer for Structural Determination Studies by Tandem PCT Pub. Date: Jun. 22, 2006 ; Rapid Communications in Mass Spectrometry, (65) Prior Publication Data vol. 9, 1227-1233 (1995). US 2009/O29.4642 A1 Dec. 3, 2009 Primary Examiner — Nicole Ippolito (74) Attorney, Agent, or Firm — Diederiks & Whitelaw, PLC Related U.S. Application Data (57) ABSTRACT A mass spectrometer is disclosed comprising a MALDI ion (60) Provisional application No. 60/641,960, filed on Jan. Source coupled to an orthogonal acceleration Time of Flight 7, 2005. mass analyzer. The mass spectrometer is operated at a first instrument setting wherein specific parent ions are selected (30) Foreign Application Priority Data by a mass filter and are accelerated to a first axial energy. The fragment ions are then orthogonally accelerated after a first Dec. 17, 2004 (GB) ...... O427632.5 delay time and first mass spectral data is obtained. The mass spectrometer is then operated at a second instrument setting (51) Int. Cl. wherein the axial energy of the parent ions is increased and HOIJ 49/40 (2006.01) the resulting fragmentions are orthogonally accelerated after (52) U.S. C. a reduced delay time. Second mass spectral data is then USPC ...... 250/287; 250/281; 250/282 obtained. The first and second mass spectral data are then (58) Field of Classification Search combined to provided a final composite . USPC ...... 250/281, 282,283, 287, 288 See application file for complete search history. 34 Claims, 3 Drawing Sheets

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U.S. Patent Aug. 13, 2013 Sheet 3 of 3 US 8,507,849 B2

SS ca SS o o SS s pus gait 00I Kisuelu KSueuI KSueu KSueui I unloads unuoads unloads UIn IOeds Gunibe dS US 8,507,849 B2 1. 2 MASS SPECTROMETER the orthogonal acceleration Time of Flight mass analyser the ions are accelerated through a constant electric field from the CROSS REFERENCE TO RELATED pusher region into the orthogonal acceleration Time of Flight APPLICATIONS flight tube. Conventional mass of the second type of This application is the National Stage of International instrument described above which comprise a MALDI ion Application No. PCT/GB2005/004911, filed on Dec. 19, Source coupled to an orthogonal acceleration Time of Flight 2005, which claims priority to and benefit of U.S. Provisional mass analyser Suffer from the problem that ions arriving at the Patent Application Ser. No. 60/641,960, filed on Jan. 7, 2005, orthogonal acceleration region of the mass analyser will have and priority to and benefit of United Kingdom Patent Appli 10 a wide range of axial energies. Accordingly, when the ions are cation No. 0427632, filed Dec. 17, 2004. The entire contents orthogonally accelerated the ion detector is only able to detect of these applications are incorporated herein by reference. and record ions having a relatively narrow or Small range of BACKGROUND OF THE INVENTION mass or mass to charge ratios. Since the orthogonal flight or 15 path length of ions in the mass analyser is limited and since The present invention relates to a mass spectrometer and a the ion detector is constrained in size then these factors (as method of mass spectrometry. will be discussed in more detail below) place a limitation on A known mass spectrometer comprises a Matrix Assisted the range of mass or mass to charge ratios of ions which can Laser Desorption Ionisation ("MALDI) coupled both be orthogonally accelerated and also Subsequently to an orthogonal acceleration Time of Flight mass analyser. detected by the ion detector of the mass analyser. Ions are orthogonally accelerated in the mass analyser and the It is therefore desired to provide an improved mass spec time of flight of the ions is measured. This enables the mass to trometer and an improved method of mass spectrometry. charge ratio of the ions to be determined. Orthogonal accel eration Time of Flight mass analysers are particularly advan SUMMARY OF THE INVENTION tageous compared to axial or in-line Time of Flight mass 25 analysers when coupled to a MALDI ion source in that the According to an aspect of the present invention there is resolution, mass calibration and mass accuracy of an orthogo provided a method of mass spectrometry comprising: nal acceleration Time of Flight mass analyser is substantially providing an orthogonal acceleration Time of Flight mass unaffected by variations in ion desorption velocities from the analyser comprising an orthogonal acceleration region; MALDI ion source. 30 providing a first packet or group of parent or precursor A further advantage of using an orthogonal acceleration ions; Time of Flight mass analyser in combination with a MALDI accelerating the first packet or group of parent or precursor ion source is that variations in the sample thickness or the ions so that the first packet or group of parent or precursor surface potential applied to the MALDI target plate do not ions possess a first axial energy; directly affect the subsequent time of flight of ions in the flight 35 fragmenting the first packet or group of parent or precursor or drift region of the orthogonal acceleration Time of Flight ions into a first plurality of fragment or daughter ions or mass analyser. allowing the first packet or group of parent or precursor ions Two different types of instrument are known. The first type to fragment into a first plurality of fragment or daughter ions; of instrument utilises a radio frequency collisional cooling orthogonally accelerating at least some of the first plurality gas cell that lowers the axial and orthogonal kinetic energy of 40 of fragment or daughter ions after a first delay time; the ions to levels appropriate for the orthogonal acceleration detecting fragment or daughter ions of the first plurality of Time of Flight mass analyser. These instruments are more fragment or daughter ions having a first range of axial ener complex, more expensive, and less efficient compared to in gies: line or axial MALDI mass spectrometers comprising a Time generating first mass spectral data relating to fragment or of Flight mass analyser. The cooling gas may promote matrix 45 daughter ions of the first plurality of fragment or daughter cluster formation that increases chemical background and ions having the first range of axial energies; reduces signal to noise. The second type of instrument does providing a second packet or group of parent or precursor not employ gaseous collisional damping and as such the ions; higher precursor ion kinetic energies permit the recording of accelerating the second packet or group of parent or pre high energy collision induced dissociation (CID) MS/MS 50 cursor ions so that the second packet or group of parent or fragmentation mass spectra. Ions are allowed to retain their precursor ions possess a second different axial energy; axial velocities and the detector of the orthogonal accelera fragmenting the second packet or group of parent or pre tion Time of Flight mass analyser has to be larger in order to cursor ions into a second plurality of fragment or daughter cope with the larger angular spread of ions caused by the large ions or allowing the second packet or group of parent or axial energy spread. One example of the second type of 55 precursor ions to fragment into a second plurality of fragment instrument is a hybrid magnetic sector orthogonal accelera or daughter ions; tion Time of Flight tandem MS/MS instrument (Bateman et orthogonally accelerating at least Some of the second plu al., Rapid Commun. Mass Spectrom. 9 (1995) 1227). The rality of fragment or daughter ions after a second delay time; instrument comprises a MALDI ion Source, a magnetic sector detecting fragment or daughter ions of the second plurality mass filter for high resolution selection of precursor ions, a 60 of fragment or daughter ions having a second range of axial collision induced dissociation (CID) gas cell and an orthogo energies: nal acceleration Time of Flight mass analyser for recording generating second mass spectral data relating to the frag the fragment or daughter ions generated in the gas cell. ment or daughter ions of the second plurality of fragment or In this instrument fragment or daughter ions retain the daughter ions having the second range of axial energies; and original parent or precursorion velocity, as such, their kinetic 65 forming a composite mass spectrum by using, combining energy is proportional to their mass. When a parent or pre or overlapping the first mass spectral data and the second cursor ion and its associated fragment or daughter ions reach mass spectral data. US 8,507,849 B2 3 4 The delay time is preferably the difference in time between (v) 15-20 us; (vi) 20-25us; (vii) 25-30 us; (viii) 30-35 us; (ix) a parent or precursor ions being generated, for example, by 35-40 us; (x) 40-45 us; (xi). 45-50 us; (xii) 50-55 us; (xiii) firing a laser at a MALDI target plate and a pusher electrode 55-60 us; (xiv) 60-65us; (xv) 65-70 us; (xvi) 70-75us; (xvii) arranged adjacent an orthogonal acceleration region of a 75-80 us; (xviii) 80-85us; (xix) 85-90 us; (XX)90-95us;(xxi) Time of Flight mass analyser being energised in order to 5 95-100 us; (xxii) 100-100 us; (xxiii) 110-120 us; (xxiv) 120 orthogonally accelerate ions into the drift or flight region of 130 us; (XXV) 130-140 us;(xxvi) 140-150 us; (xxvii) 150-160 the Time of Flight mass analyser. us; (xxviii) 160-170 us; (xxix) 170-180 us; (XXX) 180-190 us; The first range of axial energies is preferably substantially (xxxi) 190-200 us; (xxxii) 200-250 us; (xxxiii) 250-300 us; the same as the second range of axial energies. (xxxiv) 300-350 us; (XXXV) 350-400 us; (XXXvi) 400-450 us; The first delay time is preferably substantially different to 10 the second delay time. (xxxvii) 450-500 us; (xxxviii) 500-1000 us; and According to the preferred embodiment there is preferably (xxxix) >1000 us. provided a first electric field region and a first field free At least some of the first plurality of fragment or daughter region. Preferably, the first field free region is arranged down ions are preferably orthogonally accelerated so that the at stream of the first electric field region. 15 least some of the first plurality of fragment or daughter ions A second electric field region is preferably provided and a possess a first orthogonal energy. The first orthogonal energy second field free region is preferably provided. The second is preferably selected from the group consisting of: (i) <1.0 field free region is preferably arranged downstream of the keV; (ii) 1.0-1.5 keV; (iii) 1.5-2.0 keV; (iv) 2.0-2.5 keV; (v) second electric field region. 2.5-3.0 keV; (vi) 3.0-3.5 keV; (vii) 3.5-4.0 keV; (viii) 4.0-4.5 One or more electrodes are preferably arranged adjacent keV; (ix)4.5-5.0 keV; (x) 5.0–5.5 keV; (xi) 5.5-6.0 keV; (xii) the orthogonal acceleration region. 6.0-6.5 keV; (xiii) 6.5-7.0 keV; (xiv) 7.0–7.5 keV; (xv) 7.5-8.0 The step of accelerating the first packet or group of parent keV; (xvi)8.0-8.5 keV; (xvii)8.5-9.0 keV; (xviii) 9.0-9.5 keV: or precursor ions preferably comprises maintaining the first (xix) 9.5-10.0 keV; (XX) 10.0-10.5 keV; (xxi) 10.5-11.0 keV: electric field and/or the first field free region and/or the second (xxii) 11.0-11.5 keV; (xxiii) 11.5-12.0 keV; (xxiv) 12.0-12.5 electric field and/or the second field free region and/or the one 25 keV; (XXV) 12.5-13.0 keV; (xxvi) 13.0-13.5 keV; (xxvii) 13.5- or more electrodes at a first electric field strength, Voltage or 14.0 keV; (xxviii) 14.0-14.5 keV; (xxix) 14.5-15.0 keV; (XXX) potential, or Voltage or potential difference. The step of accel 15.0-15.5 keV; (xxxi) 15.5-16.0 keV; (xxxii) 16.0-16.5 keV: erating the second packet or group of parent or precursor ions (xxxiii) 16.5-17.0 keV; (xxxiv) 17.0-17.5 keV; (XXXV) 17.5- preferably comprises maintaining the first electric field and/ 18.0 keV; (XXXvi) 18.0-18.5 keV; (xxxvii) 18.5-19.0 keV: or the first field free region and/or the second electric field 30 (xxxviii) 19.0-19.5 keV; (xxxix) 19.5-20.0 keV; (xl) >20 keV. and/or the second field free region and/or the one or more The second axial energy is preferably selected from the electrodes at a second electric field strength, voltage or poten group consisting of: (i) <20 eV; (ii) 20-40 eV; (iii) 40-60 eV: tial, or voltage or potential difference. The second electric (iv) 60-80 eV; (v) 80-100 eV; (vi) 100-120 eV; (vii) 120-140 field strength, Voltage or potential, or Voltage or potential eV; (viii) 140-160 eV; (ix) 160-180 eV; (x) 180-200 eV; (xi) difference differs from the first electric field strength, Voltage 35 200-220 eV; (xii) 220-240 eV; (xiii) 240-260 eV; (xiv) 260 or potential, or voltage or potential difference by at least 1%, 280 eV; (xv) 280-300 eV; (xvi) 300-320 eV; (xvii) 320-340 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, eV; (xviii) 340-360 eV; (xix) 360-380 eV; (xx) 380-400 eV: 100%, 110%, 120%, 130%, 140%, 150%, 160%, 170%, (xxi) 400-420 eV; (xxii) 420-440 eV; (xxiii) 440-460 eV: 180%, 190%, 200%, 2.10%, 220%, 23.0%, 24.0%, 250%, (xxiv) 460-480 eV; (XXV) 480-500 eV; (xxvi) 500-550 eV: 260%, 270%. 280%, 2.90%, 300%, 350%, 400%, 450% or 40 (xxvii) 550-600 eV; (xxviii) 600-650 eV; (xxix) 650-700 eV; 500%. (xxx) 700-750 eV; (xxxi) 750-800 eV; (xxxii) 800-850 eV; According to an embodiment the first axial energy is (xxxiii) 850-900 eV; (xxxiv) 900-950 eV; (xxxv) 950-1000 selected from the group consisting of: (i) <20 eV; (ii) 20-40 eV; and (XXXVi) > 1 keV. eV; (iii) 40-60 eV; (iv) 60-80 eV; (v) 80-100 eV; (vi) 100-120 The second axial energy is preferably selected from the eV; (vii) 120-140 eV; (viii) 140-160 eV; (ix) 160-180 eV; (x) 45 group consisting of: (i) 1.0-1.2 keV; (ii) 1.2-1.4 keV; (iii) 180-200 eV; (xi) 200-220 eV; (xii) 220-240 eV; (xiii) 240-260 1.4-1.6 keV; (iv) 1..6-1.8 keV; (v) 1.8-2.0 keV; (vi) 2.0-2.2 eV; (xiv) 260-280 eV; (xv) 280-300 eV; (xvi) 300-320 eV; keV; (vii) 2.2-2.4 keV; (viii) 2.4-2.6 keV; (ix) 2.6-2.8 keV; (x) (xvii)320-340 eV; (xviii)340-360 eV; (xix)360-380 eV; (xx) 2.8-3.0 keV; (xi) 3.0-3.2 keV; (xii) 3.2-3.4 keV; (xiii) 3.4-3.6 380-400 eV; (xxi) 400-420 eV; (xxii) 420-440 eV; (xxiii) keV; (xiv) 3.6–3.8 keV; (XV) 3.8-4.0 keV; (xvii) 4.0–4.2 keV: 440-460 eV; (xxiv) 460-480 eV; (XXV) 480-500 eV; (xxvi) 50 (xvii) 4.2–4.4 keV; (xviii) 4.4–4.6 keV; (xix)4.6–4.8 keV; (xx) 500-550 eV; (xxvii) 550-600 eV; (xxviii) 600-650 eV; (xxix) 4.8–5.0 keV; (xxi) 5.0–5.5 keV; (xxii) 5.5-6.0 keV; (xxiii) 650-700 eV; (xxx) 700-750 eV; (xxxi) 750-800 eV; (xxxii) 6.0-6.5 keV; (xxiv) 6.5-7.0 keV; (XXV) 7.0–7.5 keV; (xxvi) 800-850 eV; (xxxiii) 850-900 eV; (xxxiv)900-950 eV; (xxxv) 7.5-8.0 keV; (xxvii) 8.0-8.5 keV; (xxviii) 8.5-9.0 keV; (xxix) 950-1000 eV; and (XXXvi) >1 keV. 9.0-9.5 keV; (XXX) 9.5-10.0 keV; and (xxxi) >10 keV. The first axial energy may be selected from the group 55 The second delay time is preferably selected from the consisting of: (i) 1.0-1.2 keV; (ii) 1.2-1.4 keV; (iii) 1.4-1.6 group consisting of: (i) <1 us; (ii) 1-5 us; (iii) 5-10 us; (iv) keV; (iv) 1..6-1.8 keV; (v) 1.8-2.0 keV; (vi) 2.0-2.2 keV; (vii) 10-15 us; (v) 15-20 us; (vi) 20-25 us; (vii) 25-30 us; (viii) 2.2-2.4 keV; (viii) 2.4-2.6 keV; (ix) 2.6-2.8 keV; (x) 2.8-3.0 30-35us; (ix)35-40 us;(x)40-45us; (xi)45-50 us;(xii) 50-55 keV; (xi) 3.0-3.2 keV; (xii) 3.2-3.4 keV; (xiii) 3.4-3.6 keV: us; (xiii) 55-60 us; (xiv) 60-65us; (XV) 65-70 us; (xvi) 70-75 (xiv) 3.6–3.8 keV; (XV) 3.8-4.0 keV; (xvi). 4.0-4.2 keV; (xvii) 60 us;(xvii) 75-80 us;(xviii)80-85us;(xix)85-90 us;(xx)90-95 4.2–4.4 keV; (xviii) 4.4–4.6 keV; (xix) 4.6–4.8 keV; (XX) 4.8- us; (xxi) 95-100 us; (xxii) 100-100, us; (xxiii) 110-120 us; 5.0 keV; (xxi) 5.0–5.5 keV; (xxii) 5.5-6.0 keV; (xxiii) 6.0-6.5 (xxiv) 120-130 us; (XXV) 130-140 us; (xxvi) 140-150 us; keV; (xxiv) 6.5-7.0 keV; (XXV) 7.0–7.5 keV; (xxvi) 7.5-8.0 (xxvii) 150-160 us; (xxviii) 160-170 us; (xxix) 170-180 us; keV; (xxvii) 8.0-8.5 keV; (xxviii) 8.5-9.0 keV; (xxix) 9.0-9.5 (XXX) 180-190 us; (xxxi) 190-200 us; (xxxii) 200-250 us; keV; (XXX) 9.5-10.0 keV; and (xxxi) >10 keV. 65 (xxxiii) 250-300 us; (xxxiv) 300-350 us; (XXXV) 350-400 us; The first delay time is preferably selected from the group (xxxvi) 400-450 us; (xxxvii) 450-500 us; (xxxviii) 500-1000 consisting of: (i)<1 us; (ii) 1-5us; (iii) 5-10 us; (iv) 10-15us: us; and (xxxix) >1000 us. US 8,507,849 B2 5 6 The at least some of the second plurality of fragment or (xxx) 700-750 eV; (xxxi) 750-800 eV; (xxxii) 800-850 eV; daughter ions are preferably orthogonally accelerated so that (xxxiii) 850-900 eV; (xxxiv) 900-950 eV; (xxxv) 950-1000 the at least some of the second plurality of fragment or daugh eV; and (XXXVi) > 1 keV. ter ions possess a second orthogonal energy. The second The third axial energy is preferably selected from the group orthogonal energy is preferably selected from the group con- 5 consisting of: (i) 1.0-1.2 keV; (ii) 1.2-1.4 keV; (iii) 1.4-1.6 sisting of: (i)<1.0 keV; (ii) 1.0-1.5 keV; (iii) 1.5-2.0 keV; (iv) keV; (iv) 1..6-1.8 keV; (v) 1.8-2.0 keV; (vi) 2.0-2.2 keV; (vii) 2.0-2.5 keV; (v) 2.5-3.0 keV; (vi) 3.0-3.5 keV; (vii) 3.5-4.0 2.2-2.4 keV; (viii) 2.4-2.6 keV; (ix) 2.6-2.8 keV; (x) 2.8-3.0 keV; (viii) 4.0-4.5 keV; (ix)4.5-5.0 keV; (x) 5.0–5.5 keV; (xi) keV; (xi) 3.0-3.2 keV; (xii) 3.2-3.4 keV; (xiii) 3.4-3.6 keV: 5.5-6.0 keV; (xii) 6.0–6.5 keV; (xiii) 6.5-7.0 keV; (xiv) 7.0–7.5 (xiv) 3.6–3.8 keV; (XV) 3.8-4.0 keV; (xvi). 4.0-4.2 keV; (xvii) keV; (xv) 7.5-8.0 keV; (xvi) 8.0-8.5 keV; (xvii) 8.5-9.0 keV: 10 4.2–4.4 keV; (xviii) 4.4–4.6 keV; (xix) 4.6–4.8 keV; (XX) 4.8- (xviii) 9.0-9.5 keV; (xix) 9.5-10.0 keV; (XX) 10.0-10.5 keV: 5.0 keV; (xxi) 5.0–5.5 keV; (xxii) 5.5-6.0 keV; (xxiii) 6.0–6.5 (xxi) 10.5-11.0 keV; (xxii) 11.0-11.5 keV; (xxiii) 11.5-12.0 keV; (xxiv) 6.5-7.0 keV; (XXV) 7.0–7.5 keV; (xxvi) 7.5-8.0 keV; (xxiv) 12.0-12.5 keV; (XXV) 12.5-13.0 keV; (xxvi) 13.0- keV; (xxvii) 8.0-8.5 keV; (xxviii) 8.5-9.0 keV; (xxix) 9.0-9.5 13.5 keV; (xxvii) 13.5-14.0 keV; (xxviii) 14.0-14.5 keV: 15 keV; (XXX) 9.5-10.0 keV; and (xxxi) >10 keV. (xxix) 14.5-15.0 keV; (XXX) 15.0-15.5 keV; (xxxi) 15.5-16.0 The third delay time is preferably selected from the group keV; (xxxii) 16.0-16.5 keV; (xxxiii) 16.5-17.0 keV; (xxxiv) consisting of: (i)<1 us; (ii) 1-5us; (iii) 5-10 us; (iv) 10-15us: 17.0-17.5 keV: (XXXV) 17.5-18.0 keV; (xxxvi). 18.0-18.5 keV: (v) 15-20 us; (vi) 20-25us; (vii) 25-30 us; (viii) 30-35 us; (ix) (xxxvii). 18.5-19.0 keV; (xxxviii) 19.0-19.5 keV; (xxxix) 35-40 us; (x) 40-45 us; (xi). 45-50 us; (xii) 50-55 us; (xiii) 19.5-20.0 keV; (x1) >20 keV. 55-60 us; (xiv) 60-65us; (xv) 65-70 us; (xvi) 70-75us; (xvii) According to the preferred embodiment, the method pref 75-80 us; (xviii) 80-85us; (xix) 85-90 us; (XX)90-95us;(xxi) erably further comprises: 95-100 us; (xxii) 100-100 us; (xxiii) 110-120 us; (xxiv) 120 providing a third packet or group of parent or precursor 130 us; (XXV) 130-140 us;(xxvi) 140-150 us; (xxvii) 150-160 1OnS, us; (xxviii) 160-170 us; (xxix) 170-180 us; (XXX) 180-190 us; accelerating the third packet or group of parent or precursor 25 (xxxi) 190-200 us; (xxxii) 200-250 us; (xxxiii) 250-300 us; ions so that the third packet or group of parent or precursor (xxxiv) 300-350 us; (XXXV) 350-400 us; (XXXvi) 400-450 us; ions possess a third different axial energy; (xxxvii) 450-500 us; (XXXviii) 500-1000 us; and (xxxix) fragmenting the third packet or group of parent or precur >1000 us. sor ions into a third plurality of fragment or daughter ions or The at least some of the third plurality of fragment or allowing the third packet or group of parent or precursor ions 30 daughter ions are preferably orthogonally accelerated so that the at least some of the third plurality of fragment or daughter to fragment into a third plurality of fragment or daughter ions: ions possess a third orthogonal energy. The third orthogonal orthogonally accelerating at least Some of the third plural energy is preferably selected from the group consisting of: (i) ity of fragment or daughter ions after a third delay time; <1.0 keV; (ii) 1.0-1.5 keV; (iii) 1.5-2.0 keV; (iv) 2.0-2.5 keV: detecting fragment or daughter ions of the third plurality of is (v) 2.5-3.0 keV; (vi) 3.0-3.5 keV; (vii) 3.5-4.0 keV; (viii) fragment or daughter ions having a third range of axial ener 4.0-4.5 keV; (ix) 4.5-5.0 keV; (x) 5.0–5.5 keV; (xi) 5.5-6.0 gies; and keV; (xii) 6.0–6.5 keV; (xiii) 6.5-7.0 keV; (xiv) 7.0–7.5 keV: generating third mass spectral data relating to fragment of (xv) 7.5-8.0 keV; (xvi) 8.0-8.5 keV; (xvii) 8.5-9.0 keV; (xviii) daughter ions of the third plurality of fragment or daughter 9.0-9.5 keV; (xix) 9.5-10.0 keV; (XX) 10.0-10.5 keV; (xxi) ions having the third range of axial energies. 40 10.5-11.0 keV; (xxii) 11.0-11.5 keV; (xxiii) 11.5-12.0 keV: The first, second and third ranges of axial energies are (xxiv) 12.0-12.5 keV; (XXV) 12.5-13.0 keV; (xxvi) 13.0-13.5 preferably substantially the same. The first, second and third keV; (xxvii) 13.5-14.0 keV; (xxviii) 14.0-14.5 keV; (xxix) delay times are preferably substantially different. The step of 14.5-15.0 keV; (XXX) 15.0-15.5 keV; (xxxi) 15.5-16.0 keV: accelerating the third packet or group of parent or precursor (xxxii) 16.0-16.5 keV; (xxxiii) 16.5-17.0 keV; (xxxiv) 17.0- ions preferably comprises maintaining the first electric field 45 17.5 keV; (XXXV) 17.5-18.0 keV; (xxxvi). 18.0-18.5 keV; (XXX and/or the first field free region and/or the second electric field vii). 18.5-19.0 keV; (xxxviii) 19.0-19.5 keV; (xxxix) 19.5- and/or the second field free region and/or the one or more 20.0 keV; (xl) >20 keV. electrodes at a third electric field strength, Voltage or poten The step of forming a composite mass spectrum preferably tial, or voltage or potential difference. The third electric field further comprises using, combining or overlapping the first strength, Voltage or potential, or Voltage or potential differ 50 mass spectral data, the second mass spectral data and the third ence preferably differs from the first and/or second electric mass spectral data. field strength, Voltage or potential, or Voltage or potential The method preferably further comprises: difference by at least 1%. 5%, 10%, 20%, 30%, 40%, 50%, providing a fourth packet or group of parent or precursor 60%, 70%, 80%, 90%, 100%, 110%, 120%, 130%, 140%, 1OnS, 150%, 160%, 1.70%, 180%, 190%, 200%, 210%, 220%, 55 accelerating the fourth packet or group of parent or precur 23.0%, 24.0%, 250%, 260%, 270%. 280%, 2.90%, 300%, sor ions so that the fourth packet or group of parent or pre 350%, 400%, 450% or 500%. cursor ions possess a fourth different axial energy; The third axial energy is preferably selected from the group fragmenting the fourth packet or group of parent or precur consisting of: (i) <20 eV; (ii) 20-40 eV; (iii) 40-60 eV; (iv) Sorions into a fourth plurality of fragment or daughter ions or 60-80 eV; (v) 80-100 eV; (vi) 100-120 eV; (vii) 120-140 eV; 60 allowing the fourth packet or group of parent or precursor (viii) 140-160 eV; (ix) 160-180 eV; (x) 180-200 eV; (xi) ions to fragment into a fourth plurality of fragment or daugh 200-220 eV; (xii) 220-240 eV; (xiii) 240-260 eV; (xiv) 260 ter ions; 280 eV; (xv) 280-300 eV; (xvi) 300-320 eV; (xvii)320-340 orthogonally accelerating at least Some of the fourth plu eV; (xviii) 340-360 eV; (xix) 360-380 eV; (xx) 380-400 eV: rality of fragment or daughter ions after a fourth delay time; (xxi) 400-420 eV; (xxii) 420-440 eV; (xxiii) 440-460 eV: 65 detecting fragment or daughter ions of the fourth plurality (xxiv) 460-480 eV; (XXV) 480-500 eV; (xxvi) 500-550 eV: of fragment or daughter ions having a fourth range of axial (xxvii) 550-600 eV; (xxviii) 600-650 eV; (xxix) 650-700 eV; energies; and US 8,507,849 B2 7 8 generating fourth mass spectral data relating to fragment of (xv) 7.5-8.0 keV; (xvi) 8.0-8.5 keV; (xvii) 8.5-9.0 keV; (xviii) daughter ions of the fourth plurality of fragment or daughter 9.0-9.5 keV; (xix) 9.5-10.0 keV; (XX) 10.0-10.5 keV; (xxi) ions having the fourth range of axial energies. 10.5-11.0 keV; (xxii) 11.0-11.5 keV; (xxiii) 11.5-12.0 keV: The first, second, third and fourth ranges of axial energies (xxiv) 12.0-12.5 keV; (XXV) 12.5-13.0 keV; (xxvi) 13.0-13.5 are preferably substantially the same. The first, second, third 5 keV; (xxvii) 13.5-14.0 keV; (xxviii) 14.0-14.5 keV; (xxix) and fourth delay times are preferably substantially different. 14.5-15.0 keV; (XXX) 15.0-15.5 keV; (xxxi) 15.5-16.0 keV: The step of accelerating the fourth packet or group of (xxxii) 16.0-16.5 keV; (xxxiii) 16.5-17.0 keV; (xxxiv) 17.0- parent or precursorions preferably comprises maintaining the 17.5 keV; (XXXV) 17.5-18.0 keV; (xxxvi). 18.0-18.5 keV; (XXX first electric field and/or the first field free region and/or the vii). 18.5-19.0 keV; (xxxviii) 19.0-19.5 keV; (xxxix) 19.5- second electric field and/or the second field free region and/or 10 20.0 keV; (xl) >20 keV. the one or more electrodes at a fourth electric field strength, The step of forming a composite mass spectrum preferably Voltage or potential, or Voltage or potential difference. further comprises using, combining or overlapping the first The fourth electric field strength, voltage or potential, or mass spectral data, the second mass spectral data, the third voltage or potential difference preferably differs from the first mass spectral data and the fourth mass spectral data. and/or second and/or third electric field strength, Voltage or 15 The method preferably further comprises: potential, or voltage or potential difference by at least 1%, providing a fifth packet or group of parent or precursor 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 1OnS, 100%, 110%, 120%, 130%, 140%, 150%, 160%, 170%, accelerating the fifth packet or group of parent or precursor 180%, 190%, 200%, 2.10%, 220%, 23.0%, 24.0%, 250%, ions so that the fifth packet or group of parent or precursor 260%, 270%. 280%, 2.90%, 300%, 350%, 400%, 450% or ions possess a fifth different axial energy; 500%. fragmenting the fifth packet or group of parent or precursor The fourth axial energy is preferably selected from the ions into a fifth plurality of fragment or daughter ions or group consisting of: (i) <20 eV; (ii) 20-40 eV; (iii) 40-60 eV: allowing the fifth packet or group of parent or precursor ions (iv) 60-80 eV; (v) 80-100 eV; (vi) 100-120 eV; (vii) 120-140 to fragment into a fifth plurality of fragment or daughter ions; eV; (viii) 140-160 eV; (ix) 160-180 eV; (x) 180-200 eV; (xi) 25 orthogonally accelerating at least some of the fifth plurality 200-220 eV; (xii) 220-240 eV; (xiii) 240-260 eV; (xiv) 260 of fragment or daughter ions after a fifth delay time; 280 eV; (xv) 280-300 eV; (xvi) 300-320 eV; (xvii)320-340 detecting fragment or daughter ions of the fifth plurality of eV; (xviii) 340-360 eV; (xix) 360-380 eV; (xx) 380-400 eV: fragment or daughter ions having a fifth range of axial ener (xxi) 400-420 eV; (xxii) 420-440 eV; (xxiii) 440-460 eV: gies; and (xxiv) 460-480 eV; (XXV) 480-500 eV; (xxvi) 500-550 eV: 30 generating fifth mass spectral data relating to fragment of (xxvii) 550-600 eV; (xxviii) 600-650 eV; (xxix) 650-700 eV; daughter ions of the fifth plurality of fragment or daughter (xxx) 700-750 eV; (xxxi) 750-800 eV; (xxxii) 800-850 eV; ions having the fifth range of axial energies. (xxxiii) 850-900 eV; (xxxiv) 900-950 eV; (xxxv) 950-1000 The first, second, third, fourth and fifth ranges of axial eV; and (XXXVi) >1 keV. energies are preferably Substantially the same. The first, sec The fourth axial energy may be selected from the group 35 ond, third, fourth and fifth delay times are preferably substan consisting of: (i) 1.0-1.2 keV; (ii) 1.2-1.4 keV; (iii) 1.4-1.6 tially different. keV; (iv) 1..6-1.8 keV; (v) 1.8-2.0 keV; (vi) 2.0-2.2 keV; (vii) The step of accelerating the fifth packet or group of parent 2.2-2.4 keV; (viii) 2.4-2.6 keV; (ix) 2.6-2.8 keV; (x) 2.8-3.0 or precursor ions preferably comprises maintaining the first keV; (xi) 3.0-3.2 keV; (xii) 3.2-3.4 keV; (xiii) 3.4-3.6 keV: electric field and/or the first field free region and/or the second (xiv) 3.6–3.8 keV; (XV) 3.8-4.0 keV; (xvi). 4.0-4.2 keV; (xvii) 40 electric field and/or the second field free region and/or the one 4.2–4.4 keV; (xviii) 4.4–4.6 keV; (xix) 4.6–4.8 keV; (XX) 4.8- or more electrodes at a fifth electric field strength, voltage or 5.0 keV; (xxi) 5.0–5.5 keV; (xxii) 5.5-6.0 keV; (xxiii) 6.0-6.5 potential, or Voltage or potential difference. keV; (xxiv) 6.5-7.0 keV; (XXV) 7.0–7.5 keV; (xxvi) 7.5-8.0 The fifth electric field strength, voltage or potential, or keV; (xxvii) 8.0-8.5 keV; (xxviii) 8.5-9.0 keV; (xxix) 9.0-9.5 voltage or potential difference preferably differs from the first keV; (XXX) 9.5-10.0 keV; and (xxxi) >10 keV. 45 and/or second and/or third and/or fourth electric field The fourth delay time is preferably selected from the group strength, Voltage or potential, or Voltage or potential differ consisting of: (i)<1 us; (ii) 1-5us; (iii) 5-10 us; (iv) 10-15us: ence by at least 1%. 5%, 10%, 20%, 30%, 40%, 50%, 60%, (v) 15-20 us; (vi) 20-25us; (vii) 25-30 us; (viii) 30-35 us; (ix) 70%, 80%, 90%, 100%, 110%, 120%, 130%, 140%, 150%, 35-40 us; (x) 40-45 us; (xi). 45-50 us; (xii) 50-55 us; (xiii) 160%, 1.70%, 180%, 190%, 200%, 210%, 220%, 23.0%, 55-60 us; (xiv) 60-65us; (xv) 65-70 us; (xvi) 70-75us; (xvii) 50 24.0%, 250%, 260%, 270%. 280%, 290%, 300%, 350%, 75-80 us; (xviii) 80-85us; (xix) 85-90 us; (XX)90-95us;(xxi) 400%, 450% or 500%. 95-100 us; (xxii) 100-100 us; (xxiii) 110-120 us; (xxiv) 120 The fifth axial energy is preferably selected from the group 130 us; (XXV) 130-140 us;(xxvi) 140-150 us; (xxvii) 150-160 consisting of: (i) <20 eV; (ii) 20-40 eV; (iii) 40-60 eV; (iv) us; (xxviii) 160-170 us; (xxix) 170-180 us; (XXX) 180-190 us; 60-80 eV; (v) 80-100 eV; (vi) 100-120 eV; (vii) 120-140 eV; (xxxi) 190-200 us; (xxxii) 200-250 us; (xxxiii) 250-300 us; 55 (viii) 140-160 eV; (ix) 160-180 eV; (x) 180-200 eV; (xi) (xxxiv) 300-350 us; (XXXV) 350-400 us; (XXXvi) 400-450 us; 200-220 eV; (xii) 220-240 eV; (xiii) 240-260 eV; (xiv) 260 (xxxvii) 450-500 us; (xxxviii) 500-1000 us; and 280 eV; (xv) 280-300 eV; (xvi) 300-320 eV; (xvii) 320-340 (xxxix) >1000 us. eV; (xviii) 340-360 eV; (xix) 360-380 eV; (xx) 380-400 eV: The at least some of the fourth plurality of fragment or (xxi) 400-420 eV; (xxii) 420-440 eV; (xxiii) 440-460 eV: daughter ions are preferably orthogonally accelerated so that 60 (xxiv) 460-480 eV; (XXV) 480-500 eV; (xxvi) 500-550 eV: the at least Some of the fourth plurality of fragment or daugh (xxvii) 550-600 eV; (xxviii) 600-650 eV; (xxix) 650-700 eV; ter ions possess a fourth orthogonal energy. The fourth (xxx) 700-750 eV; (xxxi) 750-800 eV; (xxxii) 800-850 eV; orthogonal energy is selected from the group consisting of: (i) (xxxiii) 850-900 eV; (xxxiv) 900-950 eV; (xxxv) 950-1000 <1.0 keV; (ii) 1.0-1.5 keV; (iii) 1.5-2.0 keV; (iv) 2.0-2.5 keV: eV; and (XXXVi) > 1 keV. (v) 2.5-3.0 keV; (vi) 3.0-3.5 keV; (vii) 3.5-4.0 keV; (viii) 65 The fifth axial energy is preferably selected from the group 4.0-4.5 keV; (ix) 4.5-5.0 keV; (x) 5.0–5.5 keV; (xi) 5.5-6.0 consisting of: (i) 1.0-1.2 keV; (ii) 1.2-1.4 keV; (iii) 1.4-1.6 keV; (xii) 6.0–6.5 keV; (xiii) 6.5-7.0 keV; (xiv) 7.0–7.5 keV: keV; (iv) 1..6-1.8 keV; (v) 1.8-2.0 keV; (vi) 2.0-2.2 keV; (vii) US 8,507,849 B2 9 10 2.2-2.4 keV; (viii) 2.4-2.6 keV; (ix) 2.6-2.8 keV; (x) 2.8-3.0 The step of accelerating the sixth packet or group of parent keV; (xi) 3.0-3.2 keV; (xii) 3.2-3.4 keV; (xiii) 3.4-3.6 keV: or precursor ions preferably comprises maintaining the first (xiv) 3.6–3.8 keV; (XV) 3.8-4.0 keV; (xvi). 4.0-4.2 keV; (xvii) electric field and/or the first field free region and/or the second 4.2–4.4 keV; (xviii) 4.4–4.6 keV; (xix) 4.6–4.8 keV; (XX) 4.8- electric field and/or the second field free region and/or the one 5.0 keV; (xxi) 5.0–5.5 keV; (xxii) 5.5-6.0 keV; (xxiii) 6.0-6.5 or more electrodes at a sixth electric field strength, Voltage or keV; (xxiv) 6.5-7.0 keV; (XXV) 7.0–7.5 keV; (xxvi) 7.5-8.0 potential, or Voltage or potential difference. keV; (xxvii) 8.0-8.5 keV; (xxviii) 8.5-9.0 keV; (xxix) 9.0-9.5 The sixth electric field strength, voltage or potential pref keV; (XXX) 9.5-10.0 keV; and (xxxi) >10 keV. erably differs from the first and/or second and/or third and/or The fifth delay time is preferably selected from the group fourth and/or fifth electric field strength, Voltage or potential, consisting of: (i)<1 us; (ii) 1-5us; (iii) 5-10 us; (iv) 10-15us: 10 or voltage or potential difference by at least 1%. 5%, 10%, (v) 15-20 us; (vi) 20-25us; (vii) 25-30 us; (viii) 30-35 us; (ix) 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 110%, 35-40 us; (x) 40-45 us; (xi). 45-50 us; (xii) 50-55 us; (xiii) 120%, 130%, 140%, 150%, 160%, 170%, 180%, 190%, 55-60 us; (xiv) 60-65us; (xv) 65-70 us; (xvi) 70-75us; (xvii) 200%, 210%, 220%, 23.0%, 240%, 250%, 260%, 270%, 75-80 us; (xviii) 80-85us; (xix) 85-90 us; (XX)90-95us;(xxi) 280%, 2.90%, 300%, 350%, 400%, 450% or 500%. 95-100 us; (xxii) 100-100 us; (xxiii) 110-120 us; (xxiv) 120 15 The sixth axial energy is preferably selected from the group 130 us; (XXV) 130-140 us;(xxvi) 140-150 us; (xxvii) 150-160 consisting of: (i) <20 eV; (ii) 20-40 eV; (iii) 40-60 eV; (iv) us; (xxviii) 160-170 us; (xxix) 170-180 us; (XXX) 180-190 us; 60-80 eV; (v) 80-100 eV; (vi) 100-120 eV; (vii) 120-140 eV; (xxxi) 190-200 us; (xxxii) 200-250 us; (xxxiii) 250-300 us; (viii) 140-160 eV; (ix) 160-180 eV; (x) 180-200 eV; (xi) (xxxiv) 300-350 us; (XXXV) 350-400 us; (XXXvi) 400-450 us; 200-220 eV; (xii) 220-240 eV; (xiii) 240-260 eV; (xiv) 260 (xxxvii) 450-500 us; (xxxviii) 500-1000 us; and 280 eV; (xv) 280-300 eV; (xvi) 300-320 eV; (xvii) 320-340 (xxxix) >1000 us. eV; (xviii) 340-360 eV; (xix) 360-380 eV; (xx) 380-400 eV: The at least some of the fifth plurality of fragment or (xxi) 400-420 eV; (xxii) 420-440 eV; (xxiii) 440-460 eV: daughter ions are preferably orthogonally accelerated so that (xxiv) 460-480 eV; (XXV) 480-500 eV; (xxvi) 500-550 eV: the at least some of the fifth plurality of fragment or daughter (xxvii) 550-600 eV; (xxviii) 600-650 eV; (xxix) 650-700 eV; ions possess a fifth orthogonal energy. The fifth orthogonal 25 (xxx) 700-750 eV; (xxxi) 750-800 eV; (xxxii) 800-850 eV; energy is preferably selected from the group consisting of: (i) (xxxiii) 850-900 eV; (xxxiv) 900-950 eV; (xxxv) 950-1000 <1.0 keV; (ii) 1.0-1.5 keV; (iii) 1.5-2.0 keV; (iv) 2.0-2.5 keV: eV; and (XXXVi) > 1 keV. (v) 2.5-3.0 keV; (vi) 3.0-3.5 keV; (vii) 3.5-4.0 keV; (viii) The sixth axial energy is preferably selected from the group 4.0-4.5 keV; (ix) 4.5-5.0 keV; (x) 5.0–5.5 keV; (xi) 5.5-6.0 consisting of: (i) 1.0-1.2 keV; (ii) 1.2-1.4 keV; (iii) 1.4-1.6 keV; (xii) 6.0–6.5 keV; (xiii) 6.5-7.0 keV; (xiv) 7.0–7.5 keV: 30 keV; (iv) 1..6-1.8 keV; (v) 1.8-2.0 keV; (vi) 2.0-2.2 keV; (vii) (XV) 7.5-8.0 keV; (xvi) 8.0-8.5 keV; (xvii) 8.5-9.0 keV; (xviii) 2.2-2.4 keV; (viii) 2.4-2.6 keV; (ix) 2.6-2.8 keV; (x) 2.8-3.0 9.0-9.5 keV; (xix) 9.5-10.0 keV; (xx) 10.0-10.5 keV; (xxi) keV; (xi) 3.0-3.2 keV; (xii) 3.2-3.4 keV; (xiii) 3.4-3.6 keV: 10.5-11.0 keV; (xxii) 11.0-11.5 keV; (xxiii) 11.5-12.0 keV: (xiv) 3.6–3.8 keV; (XV) 3.8-4.0 keV; (xvi). 4.0-4.2 keV; (xvii) (xxiv) 12.0-12.5 keV; (XXV) 12.5-13.0 keV; (xxvi) 13.0-13.5 4.2–4.4 keV; (xviii) 4.4–4.6 keV; (xix) 4.6–4.8 keV; (XX) 4.8- keV; (xxvii) 13.5-14.0 keV; (xxviii) 14.0-14.5 keV; (xxix) 35 5.0 keV; (xxi) 5.0–5.5 keV; (xxii) 5.5-6.0 keV; (xxiii) 6.0–6.5 14.5-15.0 keV; (XXX) 15.0-15.5 keV; (xxxi) 15.5-16.0 keV: keV; (xxiv) 6.5-7.0 keV; (XXV) 7.0–7.5 keV; (xxvi) 7.5-8.0 (xxxii) 16.0-16.5 keV; (xxxiii) 16.5-17.0 keV; (xxxiv) 17.0- keV; (xxvii) 8.0-8.5 keV; (xxviii) 8.5-9.0 keV; (xxix) 9.0-9.5 17.5 keV; (XXXV) 17.5-18.0 keV; (xxxvi). 18.0-18.5 keV; (XXX keV; (XXX) 9.5-10.0 keV; and (xxxi) >10 keV. vii). 18.5-19.0 keV: (XXXviii) 19.0-19.5 keV; (xxxix) 19.5- The sixth delay time is preferably selected from the group 20.0 keV; (xl) >20 keV. 40 consisting of: (i)<1 us; (ii) 1-5us; (iii) 5-10 us; (iv) 10-15us: The step of forming a composite mass spectrum preferably (v) 15-20 us; (vi) 20-25us; (vii) 25-30 us; (viii) 30-35 us; (ix) further comprises using, combining or overlapping the first 35-40 us; (x) 40-45 us; (xi). 45-50 us; (xii) 50-55 us; (xiii) mass spectral data, the second mass spectral data, the third 55-60 us; (xiv) 60-65us; (xv) 65-70 us; (xvi) 70-75us; (xvii) mass spectral data, the fourth mass spectral data and the fifth 75-80 us; (xviii) 80-85us; (xix) 85-90 us; (XX)90-95us;(xxi) mass spectral data. 45 95-100 us; (xxii) 100-100 us; (xxiii) 110-120 us; (xxiv) 120 The method preferably further comprises: 130 us; (XXV) 130-140 us;(xxvi) 140-150 us; (xxvii) 150-160 providing a sixth packet or group of parent or precursor us; (xxviii) 160-170 us; (xxix) 170-180 us; (XXX) 180-190 us; 1OnS, (xxxi) 190-200 us; (xxxii) 200-250 us; (xxxiii) 250-300 us; accelerating the sixth packet or group of parent or precur (xxxiv) 300-350 us; (XXXV) 350-400 us; (XXXvi) 400-450 us; Sorions so that the sixth packet or group of parent or precursor 50 (xxxvii) 450-500 us; (xxxviii) 500-1000 us; and ions possess a sixth different axial energy; (xxxix) >1000 us. fragmenting the sixth packet or group of parent or precur The at least some of the sixth plurality of fragment or sor ions into a sixth plurality of fragment or daughter ions or daughter ions are preferably orthogonally accelerated so that allowing the sixth packet or group of parent or precursor ions the at least some of the sixth plurality of fragment or daughter to fragment into a sixth plurality of fragment or daughter ions; 55 ions possess a sixth orthogonal energy. The sixth orthogonal orthogonally accelerating at least Some of the sixth plural energy is preferably selected from the group consisting of: (i) ity of fragment or daughter ions after a sixth delay time; <1.0 keV; (ii) 1.0-1.5 keV; (iii) 1.5-2.0 keV; (iv) 2.0-2.5 keV: detecting fragment or daughter ions of the sixth plurality of (v) 2.5-3.0 keV; (vi) 3.0-3.5 keV; (vii) 3.5-4.0 keV; (viii) fragment or daughter ions having a sixth range of axial ener 4.0-4.5 keV; (ix) 4.5-5.0 keV; (x) 5.0–5.5 keV; (xi) 5.5-6.0 gies; and 60 keV; (xii) 6.0–6.5 keV; (xiii) 6.5-7.0 keV; (xiv) 7.0–7.5 keV: generating sixth mass spectral data relating to fragment of (xv) 7.5-8.0 keV; (xvi) 8.0-8.5 keV; (xvii) 8.5-9.0 keV; (xviii) daughter ions of sixth plurality of fragment or daughter ions 9.0-9.5 keV; (xix) 9.5-10.0 keV; (XX) 10.0-10.5 keV; (xxi) having the sixth range of axial energies. 10.5-11.0 keV; (xxii) 11.0-11.5 keV; (xxiii) 11.5-12.0 keV: The first, second, third, fourth, fifth and sixth ranges of (xxiv) 12.0-12.5 keV; (XXV) 12.5-13.0 keV; (xxvi) 13.0-13.5 axial energies are preferably substantially the same. The first, 65 keV; (xxvii) 13.5-14.0 keV; (xxviii) 14.0-14.5 keV; (xxix) second, third, fourth, fifth and sixth delay times are preferably 14.5-15.0 keV; (XXX) 15.0-15.5 keV; (xxxi) 15.5-16.0 keV: substantially different. (xxxii) 16.0-16.5 keV; (xxxiii) 16.5-17.0 keV; (xxxiv) 17.0- US 8,507,849 B2 11 12 17.5 keV; (XXXV) 17.5-18.0 keV; (xxxvi). 18.0-18.5 keV; (XXX should be understood as meaning a device wherein X and Y vii). 18.5-19.0 keV: (XXXviii) 19.0-19.5 keV; (xxxix) 19.5- combine to form a product and wherein the product does not 20.0 keV; (xl) >20 keV. necessarily then fragment. The step of forming a composite mass spectrum preferably The step of allowing ions to fragment preferably comprises further comprises using, combining or overlapping the first allowing ions to fragment by Post Source Decay (“PSD') mass spectral data, the second mass spectral data, the third The method preferably further comprises providing an mass spectral data, the fourth mass spectral data, the fifth electrostatic energy analyser and/or a mass filter and/oranion mass spectral data and the sixth mass spectral data. gate for selecting specific parent or precursor ions. The mass According to an embodiment the first axial energy and/or filter preferably comprises a magnetic sector mass filter, an the second axial energy and/or the third axial energy and/or 10 the fourth axial energy and/or the fifth axial energy and/or the RF quadrupole mass filter, a Wien filter or an orthogonal sixth axial energy are preferably substantially different from acceleration Time of Flight mass filter. one another. According to an embodiment the first delay time According to another aspect of the present invention there and/or the second delay time and/or the third delay time is provided a mass spectrometer comprising: and/or the fourth delay time and/or the fifth delay time and/or 15 an orthogonal acceleration Time of Flight mass analyser the sixth delay time are preferably substantially different comprising an orthogonal acceleration region; from one another. According to an embodiment the first a control system which is arranged to: orthogonal energy and/or the second orthogonal energy and/ (i) accelerate a first packet or group of parent or precursor or the third orthogonal energy and/or the fourth orthogonal ions so that the first packet or group of parent or precursor energy and/or the fifth orthogonal energy and/or the sixth ions possesses a first axial energy; orthogonal energy are preferably Substantially the same. (ii) fragment the first packet or group of parent or precursor The method preferably further comprises providing a col ions into a first plurality of fragment or daughter ions or allow lision, fragmentation or reaction device. the first packet or group of parent or precursor ions to frag The collision, fragmentation or reaction device is prefer ment into a first plurality of fragment or daughter ions; ably arranged to fragment ions by Collisional Induced Dis 25 (iii) orthogonally accelerate at least some of the first plu sociation (“CID). rality of fragment or daughter ions after a first delay time; According to an alternative embodiment the collision, (iv) accelerate a second packet or group of parent or pre fragmentation or reaction device is selected from the group cursor ions so that the second packet or group of parent or consisting of: (i) a Surface Induced Dissociation (“SID') precursor ions possesses a second different axial energy; fragmentation device; (ii) an Electron Transfer Dissociation 30 (V) fragment the second packet or group of parent or pre fragmentation device; (iii) an Electron Capture Dissociation cursor ions into a second plurality of fragment or daughter fragmentation device; (iv) an Electron Collision or Impact ions or allowing the second packet or group of parent or Dissociation fragmentation device; (v) a Photo Induced Dis precursor ions to fragment into a second plurality of fragment sociation (“PID) fragmentation device; (vi) a Laser Induced or daughter ions; and Dissociation fragmentation device; (vii) an infrared radiation 35 (vi) orthogonally accelerate at least Some of the second induced dissociation device; (viii) an ultraviolet radiation plurality of fragment or daughter ions after a second delay induced dissociation device; (ix) a nozzle-skimmer interface time; fragmentation device; (X) an in-source fragmentation device; an ion detector which is arranged to: (xi) an ion-source Collision Induced Dissociation fragmen (i) detect fragment or daughter ions of the first plurality of tation device; (xii) a thermal or temperature source fragmen 40 fragment or daughter ions having a first range of axial ener tation device; (xiii) an electric field induced fragmentation gies: device; (xiv) a magnetic field induced fragmentation device; (ii) detect fragment or daughter ions of the second plurality (XV) an enzyme digestion or enzyme degradation fragmenta of fragment or daughter ions having a second range of axial tion device; (Xvi) an ion-ion reaction fragmentation device; energies: (Xvii) an ion-molecule reaction fragmentation device: (Xviii) 45 the mass spectrometer further comprising: an ion-atom reaction fragmentation device; (XiX) an ion means arranged to generate first mass spectral data relating metastable ion reaction fragmentation device; (XX) an ion to fragment or daughter ions of the first plurality of fragment metastable molecule reaction fragmentation device; (XXi) an or daughter ions having the first range of axial energies; ion-metastable atom reaction fragmentation device; (XXii) an means arranged to generate second mass spectral data ion-ion reaction device for reacting ions to form adduct or 50 relating to the fragment or daughter ions of the second plu product ions; (XXiii) an ion-molecule reaction device for rality of fragment or daughter ions having the second range of reacting ions to form adduct or productions; (XXiv) an ion axial energies; and atom reaction device for reacting ions to form adduct or means arranged to form a composite mass spectrum by productions; (XXV) an ion-metastable ion reaction device for using, combining or overlapping the first mass spectral data reacting ions to form adduct or productions; (XXVi) an ion 55 and the second mass spectral data. metastable molecule reaction device for reacting ions to form The first range of axial energies is preferably substantially adduct or productions; and (XXVii) an ion-metastable atom the same as the second range of axial energies. reaction device for reacting ions to form adduct or product The first delay time is preferably substantially different to ions. the second delay time. A reaction device should be understood as comprising a 60 The mass spectrometer preferably further comprises a first device wherein ions, atoms or molecules are rearranged or electric field region and a first field free region. The first field reacted so as to form a new species of ion, atom or molecule. free region is preferably arranged downstream of the first An X-Y reaction fragmentation device should be understood electric field region. as meaning a device wherein X and Y combine to form a The mass spectrometer preferably further comprises a sec product which then fragments. This is different to a fragmen 65 ond electric field region and a second field free region. The tation device perse wherein ions may be caused to fragment second field free region is preferably arranged downstream of without first forming a product. An X-Y reaction device the second electric field region. US 8,507,849 B2 13 14 The mass spectrometer preferably further comprises one or keV; (ii) 1.0-1.5 keV; (iii) 1.5-2.0 keV; (iv) 2.0-2.5 keV; (v) more electrodes arranged adjacent the orthogonal accelera 2.5-3.0 keV; (vi) 3.0-3.5 keV; (vii) 3.5-4.0 keV; (viii) 4.0-4.5 tion region. keV; (ix)4.5-5.0 keV; (x) 5.0–5.5 keV; (xi) 5.5-6.0 keV; (xii) The control system is preferably arranged to maintain the 6.0-6.5 keV; (xiii) 6.5-7.0 keV; (xiv) 7.0–7.5 keV; (xv) 7.5-8.0 first electric field and/or the first field free region and/or the 5 keV; (xvi)8.0-8.5 keV; (xvii)8.5-9.0 keV; (xviii) 9.0-9.5 keV: second electric field and/or the second field free region and/or (xix) 9.5-10.0 keV; (XX) 10.0-10.5 keV; (xxi) 10.5-11.0 keV: the one or more electrodes at a first electric field strength, Voltage or potential, or Voltage or potential difference in order (xxii) 11.0-11.5 keV; (xxiii) 11.5-12.0 keV; (xxiv) 12.0-12.5 to accelerate the first packet or group of parent or precursor keV; (XXV) 12.5-13.0 keV; (xxvi) 13.0-13.5 keV; (xxvii) 13.5- 1O.S. 10 14.0 keV; (xxviii) 14.0-14.5 keV; (xxix) 14.5-15.0 keV; (XXX) The control system is preferably arranged to maintain the 15.0-15.5 keV; (xxxi) 15.5-16.0 keV; (xxxii) 16.0-16.5 keV: first electric field and/or the first field free region and/or the (xxxiii) 16.5-17.0 keV; (xxxiv) 17.0-17.5 keV; (XXXV) 17.5- second electric field and/or the second field free region and/or 18.0 keV; (XXXvi) 18.0-18.5 keV; (xxxvii) 18.5-19.0 keV: the one or more electrodes at a second electric field strength, (xxxviii) 19.0-19.5 keV; (xxxix) 19.5-20.0 keV; (xl) >20 keV. Voltage or potential, or Voltage or potential difference in order 15 The second axial energy is preferably selected from the to accelerate the second packet or group of parent or precursor group consisting of: (i) <20 eV; (ii) 20-40 eV; (iii) 40-60 eV: 1O.S. (iv) 60-80 eV; (v) 80-100 eV; (vi) 100-120 eV; (vii) 120-140 The second electric field strength, Voltage or potential, or eV; (viii) 140-160 eV; (ix) 160-180 eV; (x) 180-200 eV; (xi) voltage or potential difference preferably differs from the first 200-220 eV; (xii) 220-240 eV; (xiii) 240-260 eV; (xiv) 260 electric field strength, Voltage or potential, or Voltage or 280 eV; (xv) 280-300 eV; (xvi) 300-320 eV; (xvii) 320-340 potential difference by at least 1%. 5%, 10%, 20%, 30%, eV; (xviii) 340-360 eV; (xix) 360-380 eV; (xx) 380-400 eV: 40%, 50%, 60%, 70%, 80%, 90%, 100%, 110%, 120%, (xxi) 400-420 eV; (xxii) 420-440 eV; (xxiii) 440-460 eV: 130%, 140%, 150%, 160%, 170%, 180%, 190%, 200%, (xxiv) 460-480 eV; (XXV) 480-500 eV; (xxvi) 500-550 eV: 210%, 220%, 23.0%, 240%, 250%, 260%, 270%. 280%, (xxvii) 550-600 eV; (xxviii) 600-650 eV; (xxix) 650-700 eV; 290%, 300%, 350%, 400%, 450% or 500%. 25 (xxx) 700-750 eV; (xxxi) 750-800 eV; (xxxii) 800-850 eV; The first axial energy is preferably selected from the group (xxxiii) 850-900 eV; (xxxiv) 900-950 eV; (xxxv) 950-1000 consisting of: (i) <20 eV; (ii) 20-40 eV; (iii) 40-60 eV; (iv) eV; and (XXXVi) > 1 keV. 60-80 eV; (v) 80-100 eV; (vi) 100-120 eV; (vii) 120-140 eV; The second axial energy is preferably selected from the (viii) 140-160 eV; (ix) 160-180 eV; (x) 180-200 eV; (xi) group consisting of: (i) 1.0-1.2 keV; (ii) 1.2-1.4 keV; (iii) 200-220 eV; (xii) 220-240 eV; (xiii) 240-260 eV; (xiv) 260 30 1.4-1.6 keV; (iv) 1..6-1.8 keV; (v) 1.8-2.0 keV; (vi) 2.0-2.2 280 eV; (xv) 280-300 eV; (xvi) 300-320 eV; (xvii)320-340 keV; (vii) 2.2-2.4 keV; (viii) 2.4-2.6 keV; (ix) 2.6-2.8 keV; (x) eV; (xviii) 340-360 eV; (xix) 360-380 eV; (xx) 380-400 eV: 2.8-3.0 keV; (xi) 3.0-3.2 keV. (xii) 3.2-3.4 keV. (xiii) 3.4-3.6 (xxi) 400-420 eV; (xxii) 420-440 eV; (xxiii) 440-460 eV: keV; (xiv) 3.6–3.8 keV; (XV) 3.8-4.0 keV; (xvii) 4.0–4.2 keV: (xxiv) 460-480 eV; (XXV) 480-500 eV; (xxvi) 500-550 eV: (xvii) 4.2–4.4 keV; (xviii) 4.4–4.6 keV; (xix)4.6–4.8 keV; (xx) (xxvii) 550-600 eV; (xxviii) 600-650 eV; (xxix) 650-700 eV; 35 4.8–5.0 keV; (xxi) 5.0–5.5 keV; (xxii) 5.5-6.0 keV; (xxiii) (xxx) 700-750 eV; (xxxi) 750-800 eV; (xxxii) 800-850 eV; 6.0-6.5 keV; (xxiv) 6.5-7.0 keV; (XXV) 7.0–7.5 keV; (xxvi) (xxxiii) 850-900 eV; (xxxiv) 900-950 eV; (xxxv) 950-1000 7.5-8.0 keV; (xxvii) 8.0-8.5 keV; (xxviii) 8.5-9.0 keV; (xxix) eV; and (XXXVi) >1 keV. 9.0-9.5 keV; (XXX) 9.5-10.0 keV; and (xxxi) >10 keV. The first axial energy is preferably selected from the group The second delay time is preferably selected from the consisting of: (i) 1.0-1.2 keV; (ii) 1.2-1.4 keV; (iii) 1.4-1.6 40 group consisting of: (i) <1 us; (ii) 1-5 us; (iii) 5-10 us; (iv) keV; (iv) 1..6-1.8 keV; (v) 1.8-2.0 keV; (vi) 2.0-2.2 keV; (vii) 10-15 us; (v) 15-20 us; (vi) 20-25 us; (vii) 25-30 us; (viii) 2.2-2.4 keV; (viii) 2.4-2.6 keV; (ix) 2.6-2.8 keV; (x) 2.8-3.0 30-35us; (ix)35-40 us;(x)40-45us; (xi)45-50 us;(xii) 50-55 keV; (xi) 3.0-3.2 keV; (xii) 3.2-3.4 keV; (xiii) 3.4-3.6 keV: us; (xiii) 55-60 us; (xiv) 60-65us; (XV) 65-70 us; (xvi) 70-75 (xiv) 3.6–3.8 keV; (XV) 3.8-4.0 keV; (xvi). 4.0-4.2 keV; (xvii) us;(xvii) 75-80 us;(xviii)80-85us;(xix)85-90 us;(xx)90-95 4.2–4.4 keV; (xviii) 4.4–4.6 keV; (xix) 4.6–4.8 keV; (XX) 4.8- 45 us; (xxi) 95-100 us; (xxii) 100-100 us; (xxiii) 110-120 us; 5.0 keV; (xxi) 5.0–5.5 keV; (xxii) 5.5-6.0 keV; (xxiii) 6.0-6.5 (xxiv) 120-130 us; (XXV) 130-140 us; (xxvi) 140-150 us; keV; (xxiv) 6.5-7.0 keV; (XXV) 7.0–7.5 keV; (xxvi) 7.5-8.0 (xxvii) 150-160 us; (xxviii) 160-170 us; (xxix) 170-180 us; keV; (xxvii) 8.0-8.5 keV; (xxviii) 8.5-9.0 keV; (xxix) 9.0-9.5 (XXX) 180-190 us; (xxxi) 190-200 us; (xxxii) 200-250 us; keV; (XXX) 9.5-10.0 keV; and (xxxi) >10 keV. (xxxiii) 250-300 us; (xxxiv) 300-350 us; (XXXV) 350-400 us; The first delay time is preferably selected from the group 50 (xxxvi) 400-450 us; (xxxvii) 450-500 us; (xxxviii) 500-1000 consisting of: (i)<1 us; (ii) 1-5us; (iii) 5-10 us; (iv) 10-15us: us; and (xxxix) >1000 us. (v) 15-20 us; (vi) 20-25us; (vii) 25-30 us; (viii) 30-35 us; (ix) The at least some of the second plurality of fragment or 35-40 us; (x) 40-45 us; (xi). 45-50 us; (xii) 50-55 us; (xiii) daughter ions are preferably orthogonally accelerated so that 55-60 us; (xiv) 60-65us; (xv) 65-70 us; (xvi) 70-75us; (xvii) the at least some of the second plurality of fragment or daugh 75-80 us; (xviii) 80-85us; (xix) 85-90 us; (XX)90-95us;(xxi) 55 ter ions possess a second orthogonal energy. The second 95-100 us; (xxii) 100-100 us; (xxiii) 110-120 us; (xxiv) 120 orthogonal energy is preferably selected from the group con 130 us; (XXV) 130-140 us;(xxvi) 140-150 us; (xxvii) 150-160 sisting of: (i)<1.0 keV; (ii) 1.0-1.5 keV; (iii) 1.5-2.0 keV; (iv) us; (xxviii) 160-170 us; (xxix) 170-180 us; (XXX) 180-190 us; 2.0-2.5 keV; (v) 2.5-3.0 keV; (vi) 3.0-3.5 keV; (vii) 3.5-4.0 (xxxi) 190-200 us; (xxxii) 200-250 us; (xxxiii) 250-300 us; keV; (viii) 4.0-4.5 keV; (ix)4.5-5.0 keV; (x) 5.0–5.5 keV; (xi) (xxxiv) 300-350 us; (XXXV) 350-400 us; (XXXvi) 400-450 us; 60 5.5-6.0 keV; (xii) 6.0–6.5 keV; (xiii) 6.5-7.0 keV; (xiv) 7.0–7.5 (xxxvii) 450-500 us; (xxxviii) 500-1000 us; and keV; (xv) 7.5-8.0 keV; (xvi) 8.0-8.5 keV; (xvii) 8.5-9.0 keV: (xxxix) >1000 us. (xviii) 9.0-9.5 keV; (xix) 9.5-10.0 keV; (XX) 10.0-10.5 keV: The at least some of the first plurality of fragment or daugh (xxi) 10.5-11.0 keV; (xxii) 11.0-11.5 keV; (xxiii) 11.5-12.0 ter ions are preferably orthogonally accelerated so that the at keV; (xxiv) 12.0-12.5 keV; (XXV) 12.5-13.0 keV; (xxvi) 13.0- least some of the first plurality of fragment or daughter ions 65 13.5 keV; (xxvii) 13.5-14.0 keV; (xxviii) 14.0-14.5 keV: possess a first orthogonal energy. The first orthogonal energy (xxix) 14.5-15.0 keV; (XXX) 15.0-15.5 keV; (xxxi) 15.5-16.0 is preferably selected from the group consisting of: (i) <1.0 keV; (xxxii) 16.0-16.5 keV; (xxxiii) 16.5-17.0 keV; (xxxiv) US 8,507,849 B2 15 16 17.0-17.5 keV: (XXXV) 17.5-18.0 keV; (xxxvi). 18.0-18.5 keV: the collision, fragmentation or reaction device with Substan (xxxvii). 18.5-19.0 keV; (xxxviii) 19.0-19.5 keV; (xxxix) tially the same Velocity and reach the orthogonal acceleration 19.5-20.0 keV; (x1) >20 keV. region at Substantially the same time. The mass spectrometerpreferably further comprises anion The mass spectrometer may comprise means arranged to Source. The ion source is preferably selected from the group cause and/or allow ions to fragment by Post Source Decay consisting of: (i) an Electrospray ionisation (ESI) ion (PSD). source; (ii) an Atmospheric Pressure Photo Ionisation The mass spectrometer may further comprise an electro (APPI) ion source; (iii) an Atmospheric Pressure Chemical static energy analyser and/or a mass filter and/or an ion gate Ionisation (APCI) ion source; (iv) a Matrix Assisted Laser for selecting specific parent or precursor ions. The mass filter Desorption Ionisation ("MALDI) ion source: (v) a Laser 10 may comprise a magnetic sector mass filter, an RF quadrupole Desorption Ionisation (“LDI) ion source: (vi) an Atmo spheric Pressure Ionisation (API) ion source: (vii) a Des mass filter, a Wien filter oran orthogonal acceleration Time of orption Ionisation on Silicon (“DIOS) ion source: (viii) an Flight mass filter. Electron Impact (“EI) ion source: (ix) a Chemical Ionisation According to another aspect of the present invention there (“CI) ion source: (X) a Field Ionisation (“FI) ion source: (xi) 15 is provided a method of mass spectrometry comprising: a (“FD) ion source; (xii) an Inductively providing an orthogonal acceleration Time of Flight mass Coupled Plasma (“ICP) ion source: (xiii) a Fast Atom Bom analyser comprising an orthogonal acceleration region; bardment (“FAB) ion source: (xiv) a Liquid Secondary Ion providing a first packet or group of parent or precursor Mass Spectrometry (“LSIMS) ion source: (XV) a Desorption ions; Electrospray Ionisation (“DESI) ion source; (xvi) a Nickel fragmenting the first packet or group of parent or precursor 63 radioactive ion source; (xvii) an Atmospheric Pressure ions into a first plurality of fragment or daughter ions or Matrix Assisted Laser Desorption Ionisation ion source; and allowing the first packet or group of parent or precursor ions (Xviii) a ion source. to fragment into a first plurality of fragment or daughter ions; The ion source may comprise a continuous or pulsed ion orthogonally accelerating at least some of the first plurality SOUC. 25 of fragment or daughter ions so that the at least some of the The mass spectrometer preferably further comprises a col first plurality of fragment or daughter ions possess a first lision, fragmentation or reaction device. orthogonal energy; The collision, fragmentation or reaction device may be detecting fragment or daughter ions of the first plurality of arranged to fragment ions by Collisional Induced Dissocia fragment or daughter ions having the first orthogonal energy; tion (“CID). 30 generating first mass spectral data relating to fragment or Alternatively, the collision, fragmentation or reaction daughter ions of the first plurality of fragment or daughter device may be selected from the group consisting of: (i) a ions having the first orthogonal energy; Surface Induced Dissociation (“SID) fragmentation device: providing a second packet or group of parent or precursor (ii) an Electron Transfer Dissociation fragmentation device; 1OnS, (iii) an Electron Capture Dissociation fragmentation device; 35 fragmenting the second packet or group of parent or pre (iv) an Electron Collision or Impact Dissociation fragmenta cursor ions into a second plurality of fragment or daughter tion device; (v) a Photo Induced Dissociation (“PID) frag ions or allowing the second packet or group of parent or mentation device; (vi) a Laser Induced Dissociation fragmen precursor ions to fragment into a second plurality of fragment tation device; (vii) an infrared radiation induced dissociation or daughter ions; device; (viii) an ultraviolet radiation induced dissociation 40 orthogonally accelerating at least Some of the second plu device; (ix) a nozzle-skimmer interface fragmentation rality of fragment or daughter ions so that the at least Some of device: (X) an in-source fragmentation device; (xi) an ion the second plurality of fragment or daughter ions possess a Source Collision Induced Dissociation fragmentation device; second different orthogonal energy; (xii) a thermal or temperature source fragmentation device; detecting fragment or daughter ions of the second plurality (xiii) an electric field induced fragmentation device; (xiv) a 45 of fragment or daughter ions having the second orthogonal magnetic field induced fragmentation device; (XV) an enzyme energy. digestion or enzyme degradation fragmentation device; (Xvi) generating second mass spectral data relating to the frag an ion-ion reaction fragmentation device; (Xvii) an ion-mol ment or daughter ions of the second plurality of fragment or ecule reaction fragmentation device: (Xviii) anion-atom reac daughter ions having the second orthogonal energy; and tion fragmentation device; (xix) an ion-metastable ion reac 50 forming a composite mass spectrum by using, combining tion fragmentation device; (XX) an ion-metastable molecule or overlapping the first mass spectral data and the second reaction fragmentation device; (XXi) an ion-metastable atom mass spectral data. reaction fragmentation device; (XXii) an ion-ion reaction The first orthogonal energy is preferably selected from the device for reacting ions to form adduct or productions; (XXiii) group consisting of: (i)<1.0 keV; (ii) 1.0-1.5 keV; (iii) 1.5-2.0 an ion-molecule reaction device for reacting ions to form 55 keV; (iv) 2.0-2.5 keV; (v) 2.5-3.0 keV; (vi) 3.0-3.5 keV; (vii) adduct or productions; (XXiv) an ion-atom reaction device for 3.5-4.0 keV; (viii) 4.0-4.5 keV; (ix) 4.5-5.0 keV; (x) 5.0–5.5 reacting ions to form adduct or productions; (XXV) an ion keV; (xi) 5.5-6.0 keV; (xii) 6.0–6.5 keV; (xiii) 6.5-7.0 keV: metastable ion reaction device for reacting ions to form (xiv) 7.0–7.5 keV; (xv) 7.5-8.0 keV; (xvi) 8.0-8.5 keV; (xvii) adduct or product ions; (XXVi) an ion-metastable molecule 8.5-9.0 keV; (xviii) 9.0-9.5 keV; (xix) 9.5-10.0 keV; (xx) reaction device for reacting ions to form adduct or product 60 10.0-10.5 keV; (xxi) 10.5-11.0 keV; (xxii) 11.0-11.5 keV: ions; and (XXVii) an ion-metastable atom reaction device for (xxiii) 11.5-12.0 keV; (xxiv) 12.0-12.5 keV; (XXV) 12.5-13.0 reacting ions to form adduct or productions. keV; (xxvi) 13.0-13.5 keV; (xxvii) 13.5-14.0 keV; (xxviii) At least some parent or precursor ions are preferably frag 14.0-14.5 keV; (xxix) 14.5-15.0 keV; (XXX) 15.0-15.5 keV: mented or reacted in use in the collision, fragmentation or (xxxi) 15.5-16.0 keV; (xxxii) 16.0-16.5 keV; (xxxiii) 16.5- reaction device to form fragment, daughter, adductor product 65 17.0 keV; (xxxiv) 17.0-17.5 keV; (XXXV) 17.5-18.0 keV: ions and wherein the fragment, daughter, adduct or product (xxxvi). 18.0-18.5 keV; (XXXvii). 18.5-19.0 keV; (xxxviii) ions and/or any corresponding parent or precursor ions exit 19.0-19.5 keV; (xxxix) 19.5-20.0 keV; (xl) >20 keV. US 8,507,849 B2 17 18 The second orthogonal energy is preferably selected from trode arranged adjacent the orthogonal acceleration region) is the group consisting of: (i) <1.0 keV; (ii) 1.0-1.5 keV; (iii) also preferably progressively decreased at each step or Sub 1.5-2.0 keV; (iv) 2.0-2.5 keV; (v) 2.5-3.0 keV; (vi) 3.0-3.5 sequent instrument setting. keV; (vii)3.5-4.0 keV; (viii) 4.0-4.5 keV; (ix)4.5-5.0 keV; (x) According to the preferred embodiment fragment or 5.0–5.5 keV; (xi) 5.5-6.0 keV; (xii) 6.0–6.5 keV; (xiii) 6.5-7.0 daughter ions having mass or mass to charge ratios within a keV; (xiv) 7.0–7.5 keV; (xv) 7.5-8.0 keV; (xvi) 8.0-8.5 keV: certain range are preferably arranged to possess appropriate (xvii) 8.5-9.0 keV; (xviii) 9.0-9.5 keV; (xix) 9.5-10.0 keV: energies such that they will follow trajectories through the (XX) 10.0-10.5 keV; (xxi) 10.5-11.0 keV; (xxii) 11.0-11.5 flight or drift region of the mass analyser and end up being keV; (xxiii) 11.5-12.0 keV; (xxiv) 12.0-12.5 keV; (XXV) 12.5- detected by the ion detector. The mass spectrometer is then 13.0 keV; (xxvi) 13.0-13.5 keV; (xxvii) 13.5-14.0 keV; (xx 10 preferably operated at second and further instrument settings and fragment or daughter ions having different masses or viii) 14.0-14.5 keV; (xxix) 14.5-15.0 keV; (XXX) 15.0-15.5 mass to charge ratios are preferably arranged to possess keV; (xxxi) 15.5-16.0 keV; (xxxii) 16.0-16.5 keV; (xxxiii) appropriate energies such that they will follow trajectories 16.5-17.0 keV; (xxxiv) 17.0-17.5 keV; (XXXV) 17.5-18.0 keV: through the flight or drift region of the mass analyser and end (xxxvi). 18.0-18.5 keV; (XXXvii). 18.5-19.0 keV; (xxxviii) 15 up being detected by the ion detector. A final composite mass 19.0-19.5 keV; (xxxix) 19.5-20.0 keV; (xl) >20 keV. spectrum is preferably produced by combining mass spectral According to another aspect of the present invention there is provided a mass spectrometer comprising: data obtained at each of the various instrument settings. an orthogonal acceleration Time of Flight mass analyser BRIEF DESCRIPTION OF THE DRAWINGS comprising an orthogonal acceleration region; a control system which is arranged to: Various embodiments of the present invention together (i) fragment a first packet or group of parent or precursor with other arrangements given for illustrative purposes only ions into a first plurality of fragment or daughter ions or allow will now be described, by way of example only, and with the first packet or group of parent or precursor ions to frag reference to the accompanying drawings in which: ment into a first plurality of fragment or daughter ions; 25 FIG. 1 shows a conventional mass spectrometer compris (ii) orthogonally accelerate at least Some of the first plural ing a MALDI ion Source coupled to an orthogonal accelera ity of fragment or daughterions so that the at least Some of the tion Time of Flight mass analyser wherein the mass spectrom first plurality of fragment or daughter ions possess a first eter further comprises a magnetic sector mass filter and a orthogonal energy; collision cell for fragmenting ions; (iii) fragment a second packet or group of parent or pre 30 FIG.2 shows a mass spectrometer according to an embodi cursor ions into a second plurality of fragment or daughter ment of the present invention comprising a MALDI ion source coupled to an orthogonal acceleration Time of Flight ions or allow the second packet or group of parent or precur mass analyser wherein the mass spectrometer further com sor ions to fragment into a second plurality of fragment or prises a first field free region and a second field free region and daughter ions; and 35 optionally a collision or fragmentation cell; and (iv) orthogonally accelerate at least Some of the second FIG. 3 shows five mass spectra acquired according to an plurality of fragment or daughter ions so that the at least some embodiment of the present invention by progressively of the second plurality of fragment or daughter ions possess a increasing the axial energy of parent or precursor ions at second different orthogonal energy; Subsequent instrument settings and by progressively reducing an ion detector which is arranged to: 40 the delay time between a pulse of ions being generated and the (i) detect fragment or daughter ions of the first plurality of pusher electrode of the Time of Flight mass analyser being fragment or daughter ions having the first orthogonal energy; energised in order to orthogonally accelerate ions into the (ii) detect fragment or daughter ions of the second plurality flight or drift region of the mass analyser. of fragment or daughter ions having the second orthogonal energy, 45 DETAILED DESCRIPTION OF THE PREFERRED the mass spectrometer further comprising: EMBODIMENTS means arranged to generate first mass spectral data relating to fragment or daughter ions of the first plurality of fragment A known mass spectrometer is shown in FIG.1. The known or daughter ions having the first orthogonal energy; mass spectrometer comprises a MALDI ion Source compris means arranged to generate second mass spectral data 50 ing a target plate 2 and laser 1. The laser 1 is arranged to emit relating to the fragment or daughter ions of the second plu a pulsed laser beam which is arranged to impinge upon the rality of fragment or daughter ions having the second target plate 2. The laser pulse causes ions to be desorbed from orthogonal energy; and the target plate 2. means arranged to form a composite mass spectrum by The MALDI ion source generates a pulse of ions which is using, combining or overlapping the first mass spectral data 55 then transmitted to a magnetic sector mass filter 3 which is and the second mass spectral data. arranged downstream of the ion source. The magnetic sector The preferred embodiment enables mass spectral data mass filter 3 comprises a high resolution mass filter which is relating to fragment or daughter ions having a wide range of arranged to mass filter parent or precursor ions emitted from mass or mass to charge ratios to be obtained without needing the ion Source Such that only parent or precursor ions having to increase the size or length of the ion detector. According to 60 a specific mass to charge ratio are onwardly transmitted by the the preferred embodiment the axial kinetic energy of parentor mass filter 3. precursor ions is preferably progressively increased in a The specific parent or precursor ions which are onwardly series of separate steps at a plurality of separate instrument transmitted by the mass filter 3 are then arranged to enter a settings. The delay time between generating a pulse of ions by Collision Induced Dissociation (“CID) gas cell 4 arranged firing the laser and orthogonally accelerating ions into the 65 downstream of the magnetic sector mass filter3. The parent or flight or drift region of the orthogonal acceleration Time of precursor ions which are transmitted by the mass filter 3 are Flight mass analyser (by applying a Voltage to a pusher elec arranged to be fragmented in the gas cell 4 such that a plural US 8,507,849 B2 19 20 ity of fragment or daughter ions are produced. The resulting wherein Mp is the mass or mass to charge ratio of a certain fragment or daughter ions are then arranged to pass from the parent or precursorion, Md is the mass or mass to charge ratio gas cell 4 to an orthogonal acceleration region of an orthogo ofa fragment or daughterion which is derived from the parent nal acceleration Time of Flight mass analyser 5. The orthogo or precursor ion, Eo is the maximum axial ion energy that an nal acceleration Time of Flight mass analyser 5 is arranged 5 ion may possess and be detected by the ion detector and EX is downstream of the gas cell 4. the orthogonal energy imparted to ions as they are orthogo The orthogonal acceleration Time of Flight mass analyser nally accelerated into the flight or drift region of the mass 5 comprises a pusher electrode 6 which is arranged adjacent analyser. the orthogonal acceleration region. Ions are arranged to ini If Md is assumed to be the lowest mass or mass to charge tially enter the mass analyser 5 along an axis 7 which passes 10 ratio fragment or daughter ion which can be detected by an through the orthogonal acceleration region. The axis 7 is also ion detector 8 having a limited length or width, then the length parallel to the plane of the pusher electrode 6. The pusher or width Ld of the ion detector 8 is given by: electrode 6 is periodically energised by applying a Voltage to the pusher electrode 6. The application of a voltage pulse to 15 the pusher electrode 6 causes an electric field in a direction (2) orthogonal to the axis 7 to be generated. The orthogonal electric field orthogonally accelerates ions present in the orthogonal acceleration region into a flight or drift region of the mass analyser 5. The flight or drift region comprises a field wherein Lx is the effective orthogonal flight or path length, free region and ions passing through the flight or drift region Eois the maximum axial ion energy that an ion may possess are arranged to become temporally separated according to and be detected by the ion detector and Ex is the orthogonal their mass to charge ratio. energy imparted to ions as they are orthogonally accelerated Anion detector 8 comprising a microchannel plate detector is arranged at the end of the flight or drift region and is into the flight or drift region of the mass analyser. arranged to detections as they arrive having passed through 25 It is apparent that the physicallength or width Ld of the ion the flight or drift region. The ion detector 8 is also arranged to detector 8 determines the lowest mass or mass to charge ratio measure the arrival time of the ions at the ion detector 8. The ion which can be detected by the ion detector 8. Accordingly, mass to charge ratio of the ions can then be derived from the it will be appreciated that the known mass spectrometer is time of flight taken for the ions to pass through the flight or only able to produce a mass spectrum of ions having a rela drift region of the mass analyser 5. 30 tively narrow or restricted range of mass or mass to charge In a mode of operation the orthogonal acceleration Time of ratios. Flight mass analyser 5 is arranged to record the mass to The orthogonal flight or path length LX is an important charge ratios of some of the fragment or daughter ions which parameter that may be maximised in order to increase mass have been produced in the gas cell 4. However, because of the resolution. However, if the orthogonal flight or path length LX limited size of the ion detector 8, the ion detector 8 is only able 35 is increased then the length of the ion detector 8 also needs to to detect fragment or daughter ions having a relatively small be increased. However, it is not practically possible to con range of masses or mass to charge ratios. tinue increasing the size or length of the ion detector 8 beyond The fragment or daughter ions produced in the gas cell 4 a certain practical limit. It will be appreciated that the cost of will retain essentially the same velocity as the parent or pre 40 an ion detector 8 increases in proportion to the size or length cursor ions from which they were derived. The kinetic energy of the ion detector 8. Furthermore, if the size or length Ld of of the fragment or daughter ions will therefore be propor the ion detector 8 is increased then it also becomes signifi tional to the mass or mass to charge ratio of the ion. cantly more difficult to maintain the necessary flatness toler In order to detect all fragment of daughter ions produced in ance for high mass resolution. Furthermore, if the length of the gas cell 4 the ion detector 8 would need to be very large or 45 the ion detector 8 were extended so that the ion detector 8 was wide since the ions which are orthogonally accelerated into the flight or drift region of the mass analyser 5 will travel able to detect relatively low mass or mass to charge ratio ions, along different trajectories which will have a large angular then the lower kinetic energies which Such ions would pos spread. The large angular spread is due to the fact that the sess is such that the ions will be more susceptible to deflection fragment or daughterions which are orthogonally accelerated or defocusing effects due to electrostatic imperfections such into the flight or drift region of the mass analyser 5 will have 50 as those resulting from unwanted Surface charging effects. a large spread of axial kinetic energies. These effects can reduce the ion transmission of low energy It can be seen from the following equation that fragment or ions and adversely effect sensitivity. daughter ions which are orthogonally accelerated into the It will be appreciated therefore that the known mass spec flight or drift region of the mass analyser 5 will follow trajec trometer suffers from the problem that it is only possible to tories which will make a wide range of differentangles C. with 55 mass analyse a relatively small proportion of the fragment or respect to the axis 7 along which ions initially entered the daughter ions which may be produced in the gas or collision mass analyser 5. The angle C. between the trajectory of a cell 4 and that it is not practical to attempt to solve this fragment or daughter ion through the flight or drift region of problem simply by making the ion detector 8 larger, wider or the mass analyser 5 and the axis 7 is shown in FIG. 1 and can 60 longer. be derived from the following relationship: FIG.2 shows a mass spectrometer according to an embodi ment of the present invention. The mass spectrometer com prises a Matrix Assisted Laser Desorption Ionisation MpEx (1) ("MALDI) ion source coupled to an orthogonal acceleration tan(a) = MdEo 65 Time of Flight mass analyser 13. Ions are preferably gener ated, released or desorbed from a target or sample plate 2 forming part of the ion source. The ions then preferably pass US 8,507,849 B2 21 22 through two separate electric field regions L. L. The electric According to an alternative embodiment, fragment or field regions L. L. may be arranged within and/or down daughter ions may be formed by Post Source Decay (“PSD') stream of the ion source. wherein the laser 1 is operated at a power such that metastable The first electric field region L is preferably arranged parent or precursor ions are formed which spontaneously immediately adjacent to the target or sample plate 2. An fragment into fragment or daughter ions after a short period of electric field is preferably maintained across the first electric time. The fragment or daughter ions will continue to pass field region L which preferably remains substantially con through the mass spectrometer at Substantially the same stant with respect to time at least until preferably substantially Velocity as their corresponding parent or precursor ions were all of the ions which have been generated pass through the travelling immediately prior to their spontaneous fragmenta first electric field region L. The electric field maintained 10 tion. Accordingly, parent or precursor ions and any corre across the first electric field region L is preferably arranged sponding fragment or daughter ions will preferably arrive at So as to accelerate parent or precursor ions to a Substantially the extraction or orthogonal acceleration region of the constant energy. The parent or precursor ions are then pref orthogonal acceleration Time of Flight mass analyser 13 at erably arranged to enter a first field free region 9 which is 15 Substantially the same time. preferably arranged downstream of the first electric field When ions arrive at the extraction or orthogonal accelera region L. tion region of the mass analyser 13, a pusher electrode 12 A second electric field region L is preferably arranged arranged preferably adjacent to the extraction or orthogonal downstream of the first electric field region L. However, acceleration region is preferably pulsed or otherwise ener according to the preferred mode of operation an electric field gised in order to extract or orthogonally accelerate ions into is not actually maintained across the second electric field the flight or drift region of the orthogonal acceleration Time region L2 although this is possible according to less preferred of Flight mass analyser 13. embodiments. A second field free region 10 is preferably The orthogonal acceleration Time of Flight mass analyser provided downstream of the second electric field region L. 13 preferably includes an ion mirror or reflectron 14 for According to the preferred embodiment the first field free 25 reflecting ions and an ion detector 15 for detecting ions. The region 9, the second electric field region L and the second reflectron or ion mirror 14 is preferably provided in order to field free region 10 may be considered as comprising a single increase the effective path length of the mass analyser 13 field free region i.e. the potential of all ion-optical compo whilst maintaining orthogonal energy focusing. The ion nents in these regions 9, L. 10 is preferably maintained detector 15 preferably comprises a microchannel plate ion Substantially the same. 30 detector although other types of ion detector may less pref The mass spectrometer preferably further comprises a erably be employed. mass filter (not shown) which is preferably arranged to select Mass spectra are preferably generated using the time of parent or precursorions having a specific mass to charge ratio. flight data recorded by the ion detector 15. In one mode of The mass filter may comprise a magnetic sector mass filter, an operation the mass spectra may include parent or precursor RF quadrupole mass filter, a Wien filter or an orthogonal 35 ions and any corresponding fragment or daughter ions pro acceleration Time of Flight mass filter. duced, for example, either by Post Source Decay or by Col The mass filter may be provided upstream of the first field lisional Induced Dissociation due to fragmentation of parent free region 9. Alternatively, the mass filter may more prefer or precursor ions within the collision or fragmentation cell 11 ably be provided in the first field free region 9, or the second or other collision, fragmentation or reaction device. electric field region L or the second field free region 10. 40 After ions have been injected into the flight or drift region Time of flight mass selection may preferably be effected by of the Time of Flight mass analyser 13, ions will arrive at the timing the flight of ions from the target plate to an orthogonal ion detector 15 at a time inversely proportional to the square extraction region (not shown) of an orthogonal acceleration root of the mass to charge ratio of the ion. A mass spectrum Time of Flight mass filter. Only ions in the vicinity of the can then be produced which may include one or more parent extraction region will be extracted or orthogonally acceler 45 or precursor ions and any corresponding fragment or daugh ated when an extraction plate (not shown) arranged adjacent ter ions created or formed either by Post Source Decay the extraction region is energised. The delay time to energise (“PSD) of the corresponding parent or precursor ions and/or the extraction region is preferably proportional to the square by Collision Induced Dissociation of corresponding parent or root of the mass or mass to charge ratio of the parent or precursor ions in the collision or fragmentation cell 11. Frag precursor ion. By default, the chosen parent or precursor ion 50 ment, daughter, product or adduct ions created by other and any associated fragment or daughter ions which travel at mechanisms in a collision, fragmentation or reaction device the same Velocity will also be extracted for mass analysis in may also be present. the orthogonal acceleration Time of Flight mass analyser The pusher electrode 12 is preferably energised when par which is preferably arranged further downstream. ent or precursor ions and/or any related fragment or daughter A collision or fragmentation cell 11 or other collision, 55 ions arrive at the orthogonal acceleration region adjacent the fragmentation or reaction device may optionally be provided pusher electrode 12. within or as part of the second field free region 10 or else The effective orthogonal path or flight length LX of ions where within the mass spectrometer. The collision or frag according to the preferred embodiment is preferably arranged mentation cell 11 may be arranged such that in a mode of So as to comprise the length of the flight or drift region from operation at least some of the ions passing through the second 60 the orthogonal acceleration region adjacent the pusher elec field free region 10 will be fragmented within the collision or trode 12 to the ion mirror 14, the effective path length within fragmentation cell 11 into fragment or daughter ions. The the ion mirror 14 and the path length from the ion mirror 14 to resulting fragment or daughter ions will then preferably pass the ion detector 15. The ion detector 15 preferably has a or continue through the remaining portion of the second field length Ld and is limited in being only able to detect ions free region 10 at substantially the same velocity as their 65 having mass to charge ratios within a particular mass to corresponding parent or precursor ions were travelling imme charge ratio range at any particular instrument setting. The diately prior to being fragmented. range of mass to charge ratios of ions which can be detected US 8,507,849 B2 23 24 at any particular instrument setting depends upon the axial wherein Mp is the mass or mass to charge ratio of the parent energies of the ions and the orthogonal energy imparted to the or precursor ion, Ep. is the axial energy of the parent or ions. precursor ion (which will now not be detected by the ion According to the preferred embodiment, in order to pro detector at the new instrument setting since the parent or duce a mass spectrum which includes fragment or daughter precursor ion will have too much kinetic energy and will ions having a wide range of mass to charge ratios, the mass therefore fly past the ion detector), Eo is the axial energy of spectrometer is preferably operated at a number of different the highest mass or mass to charge ratio ion which may be and Subsequent instrument settings and mass spectral data detected by the ion detector as the previous instrument setting and/or a separate mass spectrum is preferably obtained at and Mhis the highest mass or mass to charge ratio ion which each separate instrument setting. 10 may be detected at the new instrument setting. According to the preferred embodiment the axial kinetic If the axial energies of parent or precursor ions are energy of fragment or daughter ions is preferably effectively increased at each new instrument setting then it will be appar progressively increased by operating the mass spectrometer ent that the axial velocities of the parent or precursor ions will at a number or series of different instrument settings. The axial kinetic energy of the parent or precursor ions is prefer 15 also be increased. Likewise, since the parent or precursor ions ably progressively increased at each separate Subsequent preferably fragment in a field free region then the axial veloci instrument setting. The parent or precursor ions which frag ties of the corresponding fragment or daughter ions will also ment preferably either by Collision Induced Dissociation or be increased at the new instrument setting. by PostSource Decay into a plurality of fragment or daughter Therefore, the times of flight of ions from the sample target ions are therefore preferably arranged to possess increasingly plate 2 through the first field free region 9 and through the greater axial kinetic energies at each instrument setting. As a second field free region 10 to reach the orthogonal accelera result same species of fragment or daughter ions which are tion region adjacent the pusher electrode 12 will be reduced. formed at each Subsequent instrument setting will preferably Accordingly, according to the preferred embodiment the possess greater axial kinetic energies. delay time between a pulse of ions being generated and the The parent or precursor ions are preferably arranged to 25 pusher electrode 12 being energised in order to orthogonally fragment in either the first field free region 9 or the second accelerate ions into the flight or drift region of the mass field free region 10. According to the preferred embodiment analyser 13 is preferably correspondingly reduced at each the first and second field free regions 9,10 are preferably Subsequent new instrument setting. maintained at Substantially the same potential at each instru The shortened delay time Tp at each new instrument setting ment setting so that the first and second field free regions 9,10 30 between a pulse of ions being generated and the pusher elec act as or form a single field free region. trode 12 being energised is preferably arranged to follow the The kinetic energy of the parent or precursor ion depends following relationship: upon the product of the ionic charge of the parent or precursor ion and the acceleration Voltage applied between the target 35 i (4) plate 2 and either the first field free region 9 and/or the second Tp = To field free region 10 and/or the pusher electrode 12 in order to axially accelerate the ions. According to a less preferred embodiment the potential of the second field free region 10 and/or the pusher electrode 12 may be varied or increased at wherein To is the time of flight of parent or precursor ions each instrument setting whilst the potential of the first field 40 (having an axial energy of Eo when the mass spectrometer free region.9 may be kept constant at each instrument setting. was operated at the previous instrument setting) to pass from According to an embodiment the potential of the target the target plate 2 to the orthogonal acceleration region adja plate 2 and/or the first field free region 9 and/or the potential cent the pusher electrode 12, Mh is the highest mass or mass of the second field free region 10 and/or the potential of the to charge ratio ion which may be detected at the new instru pusher electrode 12 may be kept constant, varied, increased or 45 ment setting and Mp is the mass to charge ratio of the parent decreased at each instrument setting. or precursor ion. At any particular instrument setting ions having masses or By rearranging Equation 2 above the range of mass or mass mass to charge ratios between a low mass or mass to charge to charge ratios of ions which can be detected by the ion ratio M1 and a high mass or mass to charge ratio Mh can be detector at any particular instrument setting is given by: arranged to be detected by the ion detector 15. The highest 50 mass or mass to charge ratio ion Mh which may be detected by the ion detector 15 at any particular instrument setting can be = 1 Ex La (5) considered as possessing an axial kinetic energy Eo. Mi W. E. L. According to the preferred embodiment the axial kinetic energy of the parent or precursor ions is preferably increased 55 from one instrument setting to the next instrument setting. wherein M1 is the lowest mass to charge ratio ion which can be According to the preferred embodiment the parent or precur detected at the particular instrument setting, Mhis the highest sor ions are preferably arranged to possess an increased axial mass to charge ratio ion which can be detected at the particu kinetic energy Such that the energy of the parent or precursor lar instrument setting, EX is the orthogonal energy imparted to ion preferably increases from an energy Eo to an energy Ep 60 ions after being orthogonally accelerated into the flight or according to the following relationship: drift region of the mass analyser, Eo is the maximum axial kinetic energy of an ion which can be detected by the ion detector at the particular instrument setting, Ld is the length or width of the ion detector and LX is the effective orthogonal E. MpEo (3) P 65 flight or path length of the mass analyser. The above ratio of the minimum mass to charge ratio to the maximum mass to charge ratio of ions which can be detected US 8,507,849 B2 25 26 by the ion detector 15 at any particular instrument setting is In order to activate or energise the pusher electrode 12 at preferably a constant at any particular instrument setting the correct time, the pusher electrode delay time T1 at the since it is assumed that the orthogonal acceleration electric second instrument setting is preferably arranged to be less field and the length or width Ld of the ion detector 15 is kept than the pusher electrode delay time T0 at the first instrument COnStant. setting. The two delay times are preferably related according According to the preferred embodiment multiple separate tO: acquisitions are performed by operating the mass spectrom eter at a number of separate instrument settings. One or more mass spectra or sets of mass spectral data are preferably (7) obtained at each separate instrument setting. The various 10 T1-To...I'llMO separate mass spectra or sets of mass spectral data are then preferably combined to form a final composite mass spec trum. wherein T1 is the pusher delay time at the second instrument According to the preferred embodiment a final composite setting, T0 is the pusher delay time at the first instrument mass spectrum may be produced which includes fragment or 15 setting, M1 is the mass to charge ratio of the first specific daughter ions and which will have a significantly greater fragment or daughterion and MO is the mass to charge ratio of range of mass or mass to charge ratios compared to a mass the parent or precursor ion. spectrum which can produced using a conventional mass Generally, in order to produce a mass spectrum incorpo spectrometer. rating ions having mass to charge ratios between M0 (the In order to illustrate the preferred embodiment, a parent or mass to charge ratio of the parent or precursor ion) and M. precursor ion having a mass to charge ratio of MO may be (wherein M, is the lowest mass or mass to charge ratio frag considered. The parent or precursor ion can be considered as ment or daughter ion) and wherein the ratio M/M is con fragmenting so as to produce a number of different fragment stant at each instrument setting then the mass spectrometer or daughter ions including five specific fragment or daughter 25 should preferably be arranged to be operated at n separate and ions having different mass to charge ratios. The five specific Subsequent instrument settings. fragment or daughter ions can be considered as having mass At each instrument setting n, the parent or precursor axial to charge ratios of M1, M2, M3, M4 and M5 wherein ion energy is preferably set to E, and the pusher electrode MO>M1>M2>M3>M4>M5. For ease of illustration only, the mass to charge ratios of the parent or precursor ions and the delay time is preferably set to T- wherein: five specific fragment or daughter ions can be considered as 30 obeying the following relationship: MO/M1=M1/M2=M2/ MO (8) M3=M3AM4=M4/M5. E 1 = E0: According to the illustrative example, the mass spectrom 2-1 eter may be arranged to operate at five separate and Subse and: quent different instrument settings. 35 At the first instrument setting ions having mass to charge Tn-I =- To...I'llV V 10 (9) ratios within the range MO to M1 may be detected and recorded by the ion detector 15. At the second instrument setting the ion detector 15 can detect and record ions having 40 wherein E is the axial kinetic energy of the parent or pre mass to chargeratios within the range M1 and M2. At the third cursor ion at the nth instrument setting, E0 is the axial kinetic instrument setting the ion detector 15 can detect and record energy of the parent or precursor ion at the first instrument ions having mass to charge ratios within the range M2 and setting, MO is the mass to charge ratio of the parent or M3. At the fourth instrument setting the ion detector 15 can precursor ion, M is the highest mass to charge ratio ion detect and record ions having mass to charge ratios within the 45 which may be detected at the nth instrument setting, M., is the range M3 and M4. At the fifth instrument setting the ion lowest mass to charge ratio ion which may be detected at the detector 15 can detect and record ions having mass to charge nth instrument setting, T0 is the pusher electrode delay time at ratios within the range M4 and M5. the first instrument setting and T is the pusher electrode At the first instrument setting parent or precursor ions delay time at the nth instrument setting. having a mass to charge ratio MO are arranged to have or 50 At each separate instrument setting mass spectral data is possess an axial kinetic energy E0. preferably acquired and a mass spectrum may optionally be At the second instrument setting the axial kinetic energy of produced. the parent or precursor ions having a mass to charge ratio MO At each instrument setting the laser 1 may be fired repeat is preferably increased from an axial kinetic energy of E0 to edly so that a mass spectrum or a set of mass spectral data may a higher axial kinetic energy E1 according to the following 55 be built up or acquired from multiple acquisitions at the same relationship: instrument setting. The mass spectra or mass spectral data recorded at each of the different and Subsequent instrument settings may then E1 =- Lv.EO.' 1 (6) preferably be added together or at least overlapped so as to 60 produce a final composite mass spectrum which preferably covers a wide range of mass to charge ratios. wherein E0 is the axial kinetic energy of the parent or precur The final composite mass spectrum may be formed by Sorions at the first instrument setting, E1 is the increased axial combining the various separate mass spectra or mass spectral kinetic energy of the parent or precursor ions at the second data sets acquired at each of the different and Subsequent instrument setting, MO is the mass to charge ratio of the parent 65 instrument settings since the calibration of the orthogonal or precursor ion and M1 is the mass to charge ratio of the first acceleration Time of Flight mass analyser is preferably sub specific fragment or daughter ion. stantially independent of the axial energies of the ions when US 8,507,849 B2 27 28 they are orthogonally accelerated into the orthogonal accel quent instrument settings by varying the Voltage applied to eration region of the mass analyser 13. the pusher electrode 12. The axial ion energy of the parent or By modifying (e.g. increasing) the axial ion energies En of precursor ions may also be varied, increased or decreased at the parent or precursor ions at each Subsequent instrument Subsequent instrument settings. The pusher electrode delay setting and by modifying (e.g. shortening or reducing) the time between generating ions and energising the pusher elec pusher electrode delay time Tn between generating ions and trode 15 may also be varied, decreased or increased at subse Subsequently energising the pusher electrode 12 at each Sub quent instrument settings. sequent instrument setting and by also acquiring mass spec Some experimental results obtained according to an tral data at each instrument setting, the yield and transmission embodiment of the present invention are shown in FIG. 3. efficiency of low mass to charge ratio fragment or daughter 10 FIG. 3 shows five mass spectra which were produced or ions can be substantially enhanced compared to conventional obtained from mass spectral data which was acquired or arrangements. obtained at five separate instrument settings. The mass spec A further advantage of the preferred embodiment is that by tral data was acquired or obtained using a mass spectrometer effectively increasing the axial kinetic energy of fragment or 15 comprising a MALDI ion source coupled to an orthogonal daughter ions at each Subsequent instrument setting, the frag acceleration Time of Flight mass analyser. The mass spec ment or daughter ions become less sensitive to unwanted trometer was Substantially similar to the mass spectrometer Surface charge effects. Another advantage of increasing the shown in FIG. 2. kinetic energy at each Subsequent instrument setting is that A peptide sample of ACTH (MH+2465.2) was used in the solid divergence angle of the fragment or daughter ions is order to obtain the experimental data. ACTH peptide ions reduced. were arranged to dissociate by Post Source Decay (“PSD') The preferred embodiment preferably enables a substantial between the MALDI sample plate and the orthogonal accel increase in ion transmission to be achieved through various eration region of the Time of Flight mass analyser. fixed apertures present within the mass spectrometer. At the first instrument setting which corresponds to the first According to a less preferred embodiment the axial ener 25 mass spectrum shown in FIG. 3, the parent or precursor ions gies of the parent or precursor ions may be reduced at each were arranged to have an axial energy of 275 eV. The delay instrument setting and the pusher electrode delay time may be time between generating a pulse of ions and energising the increased at each instrument setting. pusher electrode in order to orthogonally accelerate the ions It is also contemplated that the axial energy of the parent or was set at 54.7 us. At the first instrument setting the maximum precursor ions and/or the pusher electrode delay time may be 30 mass to charge ratio of ions of interest was set at 2465 Da. varied in a non-progressive, non-linear or even random man At the second instrument setting which corresponds to the . second mass spectrum shown in FIG. 3, the parent or precur According to a less preferred embodiment instead of alter sor ions were arranged to have an axial energy of 511 eV. The ing or increasing the axial energy of the parent or precursor delay time between generating a pulse of ions and energising ions at Subsequent instrument settings, the orthogonal energy 35 the pusher electrode in order to orthogonally accelerate the imparted to the ions in the orthogonal acceleration region at ions was set at 40.0 us. At the second instrument setting the each instrument setting may be varied by altering or changing maximum mass to charge ratio of ions of interest was set at the voltage or potential applied to the pusher electrode 12 at 1327 Da. each instrument setting. At the third instrument setting which corresponds to the According to this embodiment the orthogonal energy EX, 40 third mass spectrum shown in FIG. 3, the parent or precursor imparted to ions at an nth instrument setting is preferably ions were arranged to have an axial energy of 972 eV. The related to the orthogonal energy EX imparted to ions at a delay time between generating a pulse of ions and energising previous instrument setting according to the relationship: the pusher electrode in order to orthogonally accelerate the ions was set at 28.8 us. At the third instrument setting the 45 10 maximum mass to charge ratio of ions of interest was set at Ex-1 = Ex. M-1 ( ) 698 Da. O At the fourth instrument setting which corresponds to the fourth mass spectrum shown in FIG.3, the parent or precursor wherein Ex, is the orthogonal energy imparted to ions at anth ions were arranged to have an axial energy of 2085 eV. The instrument setting, EX is the orthogonal energy imparted to 50 delay time between generating a pulse of ions and energising ions at a first or original instrument setting, M is the highest the pusher electrode in order to orthogonally accelerate the mass to charge ratio ion which may be detected at the nth ions was set at 19.4Ls. At the fourth instrument setting the instrument setting, M is the lowest mass to charge ratio ion maximum mass to charge ratio of ions of interest was set at which may be detected by the ion detector at the nth instru 325 Da. ment setting and MO is the mass to charge ratio of the parent 55 At the fifth instrument setting which corresponds to the or precursor ion. fifth mass spectrum shown in FIG. 3, the parent or precursor According to this less preferred embodiment the delay time ions were arranged to have an axial energy of 4000 eV. The between generating ions and energising the pusher electrode delay time between generating a pulse of ions and energising 12 may be kept Substantially constant from one instrument the pusher electrode in order to orthogonally accelerate the setting to the next. Further improvements to this less preferred 60 ions was set at 13.7 us. At the fifth instrument setting the embodiment are contemplated by also modifying the Voltages maximum mass to charge ratio of ions of interest was set at applied to either the electrodes forming the flight or drift 169 Da. region of the mass analyser 13 and/or the electrodes of the ion According to this particular example the orthogonal energy mirror or reflectron 14 So as to ensure that spatial time focus EX imparted to ions at each of the separate and Subsequent ing is also achieved at the ion detector 15. 65 instrument settings was kept substantially constant at 9500 According to an embodiment of the present invention the eV. The effective orthogonal flight or path length Lx was 0.8 orthogonal energy imparted to ions may be altered in Subse m and the length of the ion detector Ld was 40 cm. US 8,507,849 B2 29 30 FIG. 3 shows the five separate mass spectra which were parent or precursor ions to fragment into a second plu acquired at the five separate and Subsequent instrument set rality of fragment or daughter ions; tings. The axial energies of the parent or precursor ions and orthogonally accelerating at least some of said second plu the corresponding delay times between generating the ions rality of fragment or daughter ions after a second delay and energising the pusher electrode for each instrument set time; ting were set by generally following equations 8 and 9 as detecting fragment or daughter ions of said second plural given above. ity of fragment or daughterions having a second range of In this particular illustrative example the ratio of the high axial energies; est mass to charge ratio ion Mh to the lowest mass to charge generating second mass spectral data relating to said frag ratio ion M1 which were detected by the ion detector at each 10 instrument setting was arranged so as to be approximately ment or daughter ions of said second plurality of frag 2.1. ment or daughter ions having said second range of axial The precise ratios of the increase in the axial energy of the energies; and parent or precursor ions and the decrease in the pusher elec forming a composite mass spectrum by using, combining trode delay time varied slightly from instrument setting to 15 or overlapping said first mass spectral data and said instrument setting but in general this ratio was generally second mass spectral data. arranged to be less than 2.1 in order to allow for there to be 2. A method as claimed in claim 1, wherein said first delay Some degree of overlap between the mass spectral data time is substantially different to said second delay time. obtained or acquired at each instrument setting. This made it 3. A method as claimed in claim 1, further comprising easier to combine the mass spectral data or mass spectrum providing a first electric field region. acquired at each of the separate instrument settings So as to 4. A method as claimed in claim 3, further comprising form a final composite mass spectrum. providing a first field free region. It can be seen from the second, third, fourth and fifth mass 5. A method as claimed in claim 4, wherein said first field spectra shown in FIG.3 that progressively lower mass or mass free region is arranged downstream of said first electric field to charge fragment or daughter ions were observed at each 25 region. Subsequent instrument setting as the axial energy of the parent 6. A method as claimed in claim 5, further comprising or precursor ions was increased and the pusher electrode providing a second electric field region. delay time was reduced according to the preferred embodi 7. A method as claimed in claim 6, further comprising ment. providing a second field free region. Although the present invention has been described with 30 8. A method as claimed in claim 7, wherein said second reference to the preferred embodiments, it will be understood field free region is arranged downstream of said second elec by those skilled in the art that various changes in form and tric field region. detail may be made without departing from the scope of the 9. A method as claimed in claim 8, further comprising invention as set forth in the accompanying claims. providing one or more electrodes arranged adjacent said The invention claimed is: 35 orthogonal acceleration region. 1. A method of mass spectrometry comprising: 10. A method as claimed in claim 9, wherein said step of providing an orthogonal acceleration Time of Flight mass accelerating said first packet or group of parent or precursor analyser comprising an orthogonal acceleration region; ions comprises maintaining said one or more electrodes at a providing a first packet or group of parent or precursor first potential and wherein said step of accelerating said sec ions; 40 ond packet or group of parent or precursor ions comprises accelerating said first packet or group of parent or precur maintaining said one or more electrodes at a second potential. Sor ions with a first potential so that said first packet or 11. A method as claimed in claim 10, wherein said second group of parent or precursor ions possess a first axial potential differs from said first potential by at least 1%. 5%, energy. 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, fragmenting said first packet or group of parent or precur 45 110%, 120%, 130%, 140%, 150%, 160%, 170%, 180%, Sorions into a first plurality of fragment or daughter ions 190%, 200%, 210%, 220%, 23.0%, 24.0%, 250%, 260%, or allowing said first packet or group of parent or pre 270%. 280%, 290%, 300%, 350%, 400%, 450% or 500%. cursor ions to fragment into a first plurality of fragment 12. A method as claimed in claim 1, further comprising: or daughter ions; providing a third packet or group of parent or precursor orthogonally accelerating at least Some of said first plural 50 1OnS, ity of fragment or daughter ions after a first delay time; accelerating said third packet or group of parent or precur detecting fragment or daughter ions of said first plurality of Sor ions so that said third packet or group of parent or fragment or daughter ions having a first range of axial precursor ions possess a third different axial energy; energies: fragmenting said third packet or group of parent or precur generating first mass spectral data relating to fragment or 55 Sor ions into a third plurality of fragment or daughter daughter ions of said first plurality of fragment or daugh ions or allowing said third packet or group of parent or ter ions having said first range of axial energies; precursor ions to fragment into a third plurality of frag providing a second packet or group of parent or precursor ment or daughter ions; ions; orthogonally accelerating at least Some of said third plu accelerating said second packet or group of parent or pre 60 rality of fragment or daughter ions after a third delay cursor ions with a second potential different from the time; first potential So that said second packet or group of detecting fragment or daughter ions of said third plurality parent or precursor ions possess a second axial energy offragment or daughterions having a third range of axial different from said first axial energy; energies; and fragmenting said second packet or group of parent or pre 65 generating third mass spectral data relating to fragment of cursor ions into a second plurality of fragment or daugh daughter ions of said third plurality of fragment or ter ions or allowing said second packet or group of daughter ions having said third range of axial energies. US 8,507,849 B2 31 32 13. A method as claimed in claim 12, wherein said first, an ion detector which is arranged to: second and third ranges of axial energies are substantially the (i) detect fragment or daughter ions of said first plurality of SaC. fragment or daughter ions having a first range of axial 14. A method as claimed in claim 12, wherein said first, energies: second and third delay times are substantially different. 5 (ii) detect fragment or daughter ions of said second plural 15. A method as claimed in claim 12, wherein said step of ity of fragment or daughterions having a second range of accelerating said third packet or group of parent or precursor axial energies; ions comprises maintaining said one or more electrodes at a said mass spectrometer further comprising: third potential. means arranged to generate first mass spectral data relating 16. A method as claimed in claim 15, wherein said third 10 to fragment or daughter ions of said first plurality of potential differs from said first or second potential by at least fragment or daughter ions having said first range of axial 1%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, energies: 100%, 110%, 120%, 130%, 140%, 150%, 160%, 170%, means arranged to generate second mass spectral data 180%, 190%, 200%, 2.10%, 220%, 23.0%, 24.0%, 250%, 15 relating to said fragment or daughter ions of said second 260%, 270%. 280%, 2.90%, 300%, 350%, 400%, 450% or plurality of fragment or daughter ions having said sec 500%. ond range of axial energies; and 17. A method as claimed in claim 12, wherein said step of means arranged to form a composite mass spectrum by forming a composite mass spectrum further comprises using, using, combining or overlapping said first mass spectral combining or overlapping said first mass spectral data, said data and said second mass spectral data. second mass spectral data and said third mass spectral data. 25. A mass spectrometer as claimed in claim 24, further 18. A method as claimed in claim 1, further comprising comprising an ion source selected from the group consisting providing a collision, fragmentation or reaction device. of: (i) an Electrospray ionisation (“EST) ion source; (ii) an 19. A method as claimed in claim 18, wherein said colli Atmospheric Pressure Photo Ionisation (APPI) ion source: Sion, fragmentation or reaction device is arranged to fragment 25 (iii) an Atmospheric Pressure Chemical Ionisation (APCI) ions by Collisional Induced Dissociation (“CID). ion source; (iv) a Matrix Assisted Laser Desorption Ionisation 20. A method as claimed in claim 1, wherein said step of ("MALDI) ion source: (v) a Laser Desorption Ionisation allowing ions to fragment comprises allowing ions to frag (“LDI) ion source: (vi) an Atmospheric Pressure Ionisation ment by Post Source Decay (“PSD). (API) ion source: (vii) a Desorption Ionisation on Silicon 21. A method as claimed in claim 1, further comprising 30 (“DIOS) ion source: (viii) an Electron Impact (“EI) ion providing an electrostatic energy analyser or a mass filter or source: (ix) a Chemical Ionisation (“CI) ion source; (x) a an ion gate for selecting specific parent or precursor ions. Field Ionisation (“FI) ion source: (xi) a Field Desorption 22. A method as claimed in claim 21, wherein said mass (“FD) ion source; (xii) an Inductively Coupled Plasma filter comprises a magnetic sector mass filter, an RF quadru (“ICP”)ion source: (xiii) a (“FAB) pole mass filter, a Wien filter or an orthogonal acceleration 35 ion source; (xiv) a Liquid Secondary Ion Mass Spectrometry Time of Flight mass filter. (“LSIMS) ion source: (XV) a Desorption Electrospray Ioni 23. A method as claimed in claim 1, wherein said first range sation (“DESI) ion source: (xvi) a Nickel-63 radioactive ion of axial energies is Substantially different from said second source: (xvii) an Atmospheric Pressure Matrix Assisted Laser range of axial energies. Desorption Ionisation ion source; and (Xviii) a Thermospray 24. A mass spectrometer comprising: 40 ion source. an orthogonal acceleration Time of Flight mass analyser 26. A mass spectrometer as claimed in claim 24, further comprising an orthogonal acceleration region; comprising a collision, fragmentation or reaction device. a control system which is arranged to: 27. A mass spectrometeras claimed in claim 26, wherein at (i) accelerate a first packet or group of parent or precursor least some parent or precursor ions are fragmented or reacted ions with a first potential so that said first packet or group 45 in use in said collision, fragmentation or reaction device to of parent or precursor ions possesses a first axial energy; form fragment, daughter, adduct or productions and wherein (ii) fragment said first packet or group of parent or precur said fragment, daughter, adduct or product ions exit said Sorions into a first plurality of fragment or daughter ions collision, fragmentation or reaction device with Substantially or allow said first packet or group of parent or precursor the same Velocity and reach said orthogonal acceleration ions to fragment into a first plurality of fragment or 50 region at Substantially the same time. daughter ions; 28. A mass spectrometer as claimed in claim 24, wherein (iii) orthogonally accelerate at least some of said first plu said first range of axial energies is substantially different from rality of fragment or daughter ions after a first delay said second range of axial energies. time; 29. A method of mass spectrometry comprising: (iv) accelerate a second packet or group of parent or pre 55 providing an orthogonal acceleration Time of Flight mass cursor ions with a second potential different from the analyser comprising an orthogonal acceleration region; first potential So that said second packet or group of providing a first packet or group of parent or precursor parent or precursor ions possesses a second axial energy ions; different from said first axial energy; fragmenting said first packet or group of parent or precur (V) fragment said second packet or group of parent or 60 Sorions into a first plurality of fragment or daughter ions precursor ions into a second plurality of fragment or or allowing said first packet or group of parent or pre daughter ions or allowing said second packet or group of cursor ions to fragment into a first plurality of fragment parent or precursor ions to fragment into a second plu or daughter ions; rality of fragment or daughter ions; and orthogonally accelerating at least some of said first plural (vi) orthogonally accelerate at least some of said second 65 ity of fragment or daughter ions with a first potential So plurality of fragment or daughter ions after a second that said at least Some of said first plurality of fragment delay time; or daughter ions possess a first orthogonal energy; US 8,507,849 B2 33 34 detecting fragment or daughter ions of said first plurality of 32. A method as claimed in claim29, wherein orthogonally fragment or daughter ions having said first orthogonal accelerating at least some of said first plurality of fragment or energy; daughter ions includes applying a first voltage and orthogo generating first mass spectral data relating to fragment or nally accelerating at least some of said second plurality of 5 fragment or daughter ions includes applying a second voltage daughter ions of said first plurality of fragment or daugh different from the first voltage so as to cause said at least some ter ions having said first orthogonal energy; of said second plurality of fragment or daughter ions possess providing a second packet or group of parent or precursor said second orthogonal energy different from said first 1Ons; orthogonal energy. fragmenting said second packet or group of parent or pre 33. A mass spectrometer comprising: cursor ions into a second plurality of fragment or daugh 10 an orthogonal acceleration Time of Flight mass analyser ter ions or allowing said second packet or group of comprising an orthogonal acceleration region; parent or precursor ions to fragment into a second plu a control system which is arranged to: rality of fragment or daughter ions; (i) fragment a first packet or group of parent or precursor orthogonally accelerating at least some of said second plu ions into a first plurality of fragment or daughter ions or rality of fragment or daughter ions with a second poten 15 allow said first packet or group of parent or precursor tial different from the first potential so that said at least ions to fragment into a first plurality of fragment or Some of said second plurality of fragment or daughter daughter ions; ions possess a second orthogonal energy different from (ii) orthogonally accelerate at least some of said first plu said first orthogonal energy; rality of fragment or daughter ions with a first potential detecting fragment or daughter ions of said second plural So that said at least some of said first plurality of frag ity of fragment or daughter ions having said second ment or daughter ions possess a first orthogonal energy; orthogonal energy; (iii) fragment a second packet or group of parent or pre generating second mass spectral data relating to said frag cursor ions into a second plurality of fragment or daugh ment or daughter ions of said second plurality of frag terions or allow said second packet or group of parent or ment or daughter ions having said second orthogonal 25 precursor ions to fragment into a second plurality of energy; and fragment or daughter ions; and (iv) orthogonally accelerate at least some of said second forming a composite mass spectrum by using, combining plurality of fragment or daughter ions with a second or overlapping said first mass spectral data and said potential different from the first potential so that said at Second mass spectral data. 30 least some of said second plurality of fragment or daugh 30. A method as claimed in claim 29, wherein said first ter ions possess a second orthogonal energy different orthogonal energy is selected from the group consisting of: (i) from said first orthogonal energy; <1.0 keV; (ii) 1.0-1.5 keV; (iii) 1.5-2.0 keV; (iv) 2.0-2.5 keV; an ion detector which is arranged to: (v) 2.5-3.0 keV; (vi) 3.0-3.5 keV; (vii) 3.5-4.0 keV; (viii) (i) detect fragment or daughter ions of said first plurality of 4.0-4.5 keV; (ix) 4.5-5.0 keV; (x) 5.0–5.5 keV; (xi) 5.5-6.0 35 fragment or daughter ions having said first orthogonal keV; (xii) 6.0–6.5 keV; (xiii) 6.5-7.0 keV; (xiv) 7.0–7.5 key: energy: (XV) 7.5-8.0 keV; (xvi) 8.0-8.5 keV; (xvii) 8.5-9.0 keV; (xviii) (ii) detect fragment or daughter ions of said second plural 9.0-9.5 keV; (xix) 9.5-10.0 keV; (xx) 10.0-10.5 keV; (xxi) ity of fragment or daughter ions having said second 10.5-11.0 keV; (xxii) 11.0-11.5 keV; (xxiii) 11.5-12.0 keV; orthogonal energy; (xxiv) 12.0-12.5 keV; (xxv) 12.5-13.0 key; (xxvi) 13.0-13.5 40 said mass spectrometer further comprising: keV; (xxvii) 13.5-14.0 keV; (xxviii) 14.0-14.5 keV; (xxix) means arranged to generate first mass spectral data relating 14.5-15.0 keV; (xxx) 15.0-15.5 keV; (xxxi) 15.5-16.0 keV; to fragment or daughter ions of said first plurality of (xxxii) 16.0-16.5 keV; (xxxiii) 16.5-17.0 keV; (xxxiv) 17.0- fragment or daughter ions having said first orthogonal 17.5 keV; (XXXV) 17.5-18.0 keV; (xxxvi). 18.0-18.5 keV; (xxx energy: vii) 18.5-19.0 key; (xxxviii) 19.0-19.5 keV; (xxxix) 19.5-20.0 45 means arranged to generate second mass spectral data keV; (xl) >20 keV. relating to said fragment or daughter ions of said second 31. A method as claimed in claim 30, wherein said second plurality of fragment or daughter ions having said sec orthogonal energy is selected from the group consisting of: (i) ond orthogonal energy; and <1.0 keV; (ii) 1.0-1.5 key; (iii) 1.5-2.0 keV; (iv) 2.0-2.5 keV; means arranged to form a composite mass spectrum by (v) 2.5-3.0 key; (vi) 3.0-3.5 keV; (vii) 3.5-4.0 keV; (viii) 50 using, combining or overlapping said first mass spectral 4.0-4.5 keV; (ix) 4.5-5.0 keV; (x) 5.0–5.5 keV; (xi) 5.5-6.0 data and said second mass spectral data. keV; (xii) 6.0–6.5 keV; (xiii) 6.5-7.0 keV; (xiv) 7.0–7.5 keV; 34. A mass spectrometer as claimed in claim 33, wherein (XV) 7.5-8.0 keV; (xvi) 8.0-8.5 key; (xvii) 8.5-9.0 keV; (xviii) the control system is further configured to orthogonally accel 9.0-9.5 keV; (xix) 9.5-10.0 keV; (xx) 10.0-10.5 keV; (xxi) erate at least some of said first plurality of fragment or daugh 10.5-11.0 keV; (xxii) 11.0-11.5 keV; (xxiii) 11.5-12.0 keV; 55 ter ions by applying a first voltage and orthogonally acceler (xxiv) 12.0-12.5 keV; (xxv) 12.5-13.0 keV; (xxvi) 13.0-13.5 ate at least some of said second plurality of fragment or keV; (xxvii) 13.5-14.0 keV; (xxviii) 14.0-14.5 keV; (xxix) daughter ions by applying a second voltage different from the 14.5-15.0 keV; (xxx) 15.0-15.5 keV; (xxxi) 15.5-16.0 keV; first Voltage so as to cause said at least some of said second (xxxii) 16.0-16.5 keV; (xxxiii) 16.5-17.0 keV; (xxxiv) 17.0- plurality of fragment or daughter ions possess said second 17.5 keV; (XXXV) 17.5-18.0 keV; (xxxvi). 18.0-18.5 keV; (xxx 60 vii). 18.5-19.0 keV; (xxxviii) 19.0-19.5 keV; (xxxix) 19.5- orthogonal energy different from said first orthogonal energy. 20.0 keV; (xl) >20 keV. ck ck ck ck ck