1. Proteomics Informatics – Overview of Mass spectrometry (Week 2) Ion Source Mass Analyzer Detector mass/charge intensity

2. Peptide Fragmentation Mass Analyzer 1 Frag- mentation DetectorIon Source Mass Analyzer 2 b y

3. Liquid Chromatography (LC)-MS/MS Mass Analyzer 1 Frag- mentation Detector intensity mass/charge Ion Source Mass Analyzer 2 LC intensity mass/charge intensity mass/charge intensity mass/charge intensity mass/charge intensity mass/charge Time intensity mass/charge intensity mass/charge intensity mass/charge intensity mass/charge intensity mass/charge intensity mass/charge intensity mass/charge intensity mass/charge intensity mass/charge

4. Ion Sources Ion Source Mass Analyzer Detector MALDI ESI mass/charge intensity

5. Electrospray Mass Analyzer Detector

6. Matrix Assisted Laser Desorption Ionization (MALDI) alpha-cyano-4-hydroxycinnamic acid Matrix Laser Mass Analyzer Detector

7. Detectors Ion Source Mass Analyzer Detector mass/charge intensity Electron Multiplier Image Current

8. Electron Multiplier Detector

9. Image Current Detector

10. Mass Analyzers Ion Source Mass Analyzer Detector Time-of-Flight Magnetic Sector Quadrupole Ion Trap (3D, linear) FTICR Orbitrap mass/charge intensity

11. Mass Spectrometry (MS)

12. Time-of-Flight Mass Spectrometry Ion Source Mass Analyzer Detector Time-of-Flight Detector Ion mirror

13. Magnetic Sector

14. Quadrupole Mass Filter y x

15. Ion Trap

16. Fourier transform ion cyclotron resonance

17. Orbitrap

18. Quadrupole Time-of-Flight Mass Spectrometer Video Mass Analyzer 1 Frag- mentation Detector Ion Source Mass Analyzer 2 CAD – Collision Activated Dissociation Quadrupole Time-of-Flight

19. Triple Quadrupole Mass Analyzer 1 Frag- mentation Detector mass/charge intensity Ion Source Mass Analyzer 2 CAD – Collision Activated Dissociation Quadrupole time m/z time m/z time m/z time m/z time m/z time m/z NO YES time m/z time m/z YES m/z time  m/z is constant

20. Linear Ion Trap / Orbitrap Mass Analyzer 1 Frag- mentation Detector Ion Source Mass Analyzer 2 ESILinear Ion Trap Orbitrap CAD ETD Olsen J V et al. Mol Cell Proteomics 2009;8:2759-2769 Frag- mentation HCD Detector Video

21. Data Independent Acquisistion mass/charge intensity 1. MS 2. MS/MS 1 3. MS/MS 2 4. MS/MS 3 mass/charge intensity mass/charge intensity mass/charge intensity mass/charge intensity 5. MS 6. MS/MS 1 7. MS/MS 2 8. MS/MS 3 9. MS 10. MS/MS 1 11. MS/MS 2 12. MS/MS 3 13. MS 14. MS/MS 1 15. MS/MS 2 16. MS/MS 3 17. MS 18. MS/MS 1 19. MS/MS 2 20. MS/MS 3 21. MS 22. MS/MS 1 23. MS/MS 2 24. MS/MS 3 …

22. 1.MS 2.MS/MS 1 3.MS/MS 2 4.MS/MS 3 5.MS/MS 4 6.MS/MS 5 7.MS/MS 6 8.MS/MS 7 9.MS/MS 8 10.MS/MS 9 11.MS/MS 10 Data Dependent Acquisistion mass/charge intensity mass/charge intensity 12. MS 13. MS/MS 1 14. MS/MS 2 15. MS/MS 3 16. MS/MS 4 17. MS/MS 5 18. MS/MS 6 19. MS/MS 7 20. MS/MS 8 21. MS/MS 9 22. MS/MS 10 …

23. Vacuum System Ion Source Mass Analyzer Detector Vacuum System Atmosphere

24. Mass Spectrometry Data Ion Source Mass Analyzer Detector mass/charge intensity

25. Mass Spectrometry Data Dimensions: Time Peptide m/z Peptide Intensity Petide fragment m/z Peptide fragment intensity...

26. Example data – MALDI-TOF Peptide intensity vs m/z

27. Fragment intensity vs m/z Example data – ESI-LC-MS/MS Time m/z MS/MS Peptide intensity vs m/z vs time

28. Slice - Scalable Data Sharing for Remote Mass Informatics Most mass spectrometry data is acquired in discovery mode, meaning that the data is amenable to open-ended analysis as our understanding of the target biochemistry increases. In this sense, mass spectrometry based discovery work is more akin to an astronomical survey, where the full list of object-types being imaged has not yet been fully elucidated, as opposed to e.g. micro-array work, where the list of probes spotted onto the slide is finite and well understood. slice.ionomix.com Developed by Manor Askenazi

29. Example Mass Spectrometry Data fenyolab.ionomix.com

30. Example Mass Spectrometry Data Peptide intensity vs time For 737.707 m/z which corresponds to 3+ of LGEHNIDVLEGNEQFINAAK

31. Example Mass Spectrometry Data Fragment intensity vs m/z For 3+ of LGEHNIDVLEGNEQFINAAK

32. Example Mass Spectrometry Data Fragment intensity vs m/z For 3+ of LGEHNIDVLEGNEQFINAAK

33. Example Mass Spectrometry Data Peptide intensity vs m/z 3+ of LGEHNIDVLEGNEQFINAAK

34. Example Mass Spectrometry Data Peptide intensity vs m/z 3+ of LGEHNIDVLEGNEQFINAAK

35. Example Mass Spectrometry Data Peptide intensity vs m/z Peptide intensity vs time

36. Example Mass Spectrometry Data ASTHTDSSAQTVSLEDYVSR 3+ in E. coli DTTTIIDGVGEEAAIQGR 2+ in E. coli ATGTSEMAPALVAAFGGK 2+ in E. coli FVPDTQAPLGIR 2+ in E. coli

37. Proteomics Informatics – Overview of Mass spectrometry (Week 2) Ion Source Mass Analyzer Detector mass/charge intensity