introduction to the crl mass spectrometry research facility · mass spectrometry research facility...
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DEPARTMENT OF CHEMISTRYMASS SPECTROMETRY RESEARCH FACILITY
Professor James McCullagh, Head of the Mass Spectrometry Research Facility
INTRODUCTION TO THE CRL MASS SPECTROMETRY RESEARCH FACILITY
October 5, 2015Mass Spectrometry website: http://www.chem.ox.ac.uk/spectroscopy/mass-spec/ Page 2
October 5, 2015Mass Spectrometry website: http://www.chem.ox.ac.uk/spectroscopy/mass-spec/ Page 3
Introduction
1. MS overview2. People3. Facilities4. Instrument locations5. Open access MS6. Trained users MS7. Project work8. Software and data storage9. Lectures and events
Mass as an inherent component of all matter
Proton = 1.0073 Neutron = 1.0087Electron = 0.00055
Mass Spectrometry website: http://www.chem.ox.ac.uk/spectroscopy/mass-spec/
Introduction to mass spectrometry
October 5, 2015Mass Spectrometry website: http://www.chem.ox.ac.uk/spectroscopy/mass-spec/ Page 5
Identification Elemental composition, chemical formula predictions, fragmentation pattern fingerprint.
Structural studies – fragmentation studies using MS can lead to information about compound structure – used in conjunction with NMR…
Quantification Determine the amounts of a compound present. LC/MS; GC/MS
Mass Spectrometry Research Facility
October 5, 2015Mass Spectrometry website: http://www.chem.ox.ac.uk/spectroscopy/mass-spec/ Page 6
Prof James McCullaghHead of Research facility
Dr James WickensMS Services Manager
Mr Colin sparrowResearch Technician
Ms Elisabete PiresResearch Associate
18 MS systems5 full time staffMultiple MS services
MS facility and staff support Research in the CRL (Part II, DPhil, Postdoc and PIs).
Dr Ashley ShepherdResearch Associate
Location of MS labs in CRL
October 5, 2015Mass Spectrometry website: http://www.chem.ox.ac.uk/spectroscopy/mass-spec/ Page 7
Basement Mass Spec Lab 2
Basement Mass Spec Lab 1
EP1
Slide 7
EP1 Mc Cullagh correct from McCulloughtElisabete Pires, 22/09/2015
Types of access to mass spectrometry
1. OPEN ACCESS MASS SPECTROMTRY walk up Instruments on ground floor (initial registration and training 10 mins.) anyone in Chemistry Dept can use.
2. MASS SPECTROMETRY SUBMISSION SERVICES: Accurate Mass Service (Lecture on this will be held in CRL in Oct)Alternative ionisation MS Proteomics service.Special samples – we can often help with samples which fall outside the remit above – please come and talk to a member of staff.
October 5, 2015
Mass Spectrometry website: http://www.chem.ox.ac.uk/spectroscopy/mass-spec/Page 8
Dr James Wickens: Mass Spectrometry Services Manager
Types of access to mass spectrometry
3. TRAINED USER INSTRUMENTS 9 instruments – require training course – can then book and use whenever you like. Open to all but please email or come and see James Wickens or James McCullagh to discuss needs first. LC/MS, GC/MS and MALDI available.
4. RESEARCH COLLABORATIONSMethod development for bespoke systemsProteomics and post-translations modifications analysisMetabolite identification
October 5, 2015
Mass Spectrometry website: http://www.chem.ox.ac.uk/spectroscopy/mass-spec/Page 9
Dr James Wickens: Mass Spectrometry Services Manager
1. Open Access Mass Spectrometry
SMALL MOLECULES (100-2000 Da)1. Runs 60,000 samples/yr2. 2 mins per sample3. Used almost continuously4. 10min familiarisation training Mon/Tue/Wed5. Sign up sheet required after NMR lecture.
PROTEIN OPEN ACCESS1. Fast column chromatography LC/MS2. 10mins per sample3. Fewer users – see a member of MS staff
for training
October 5, 2015Mass Spectrometry website: http://www.chem.ox.ac.uk/spectroscopy/mass-spec/ Page 10
Small molecule open access
Protein open access
Ground floor MS room
Ground floor MS room
Where: Ground floor mass spec room
October 5, 2015Mass Spectrometry website: http://www.chem.ox.ac.uk/spectroscopy/mass-spec/ Page 11
Open access: Results
October 5, 2015
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Small molecule open access Protein open access
Small moleculesResults are emailed as a PDFRaw data files available from2am next day on thechemistry network: \\FILES-SUPPORT\Mass Spectrometry\Open Access
Protein Open AccessResults deposited as PDF in sepárate folder in samelocation.
Results: PDF emailed
\\FILES-SUPPORT\Mass Spectrometry\Open Access
Results: PDF emailed
Adducts M+HM+NaM-H
2. MS servicesSmall Molecules (organic and inorganic compounds):1. Accurate Mass Service (novel compounds)2. High resolution Service (unable to obtain results by open
access)
Biological mass spectrometry:1. Proteomics Service2. Metabolomics
October 5, 2015Mass Spectrometry website: http://www.chem.ox.ac.uk/spectroscopy/mass-spec/ Page 13
Small molecules sample login: Ground floor Lab 20.062 (same as open access)
Biological MS services: Submit samples to a member of MS staff in basement
Small molecule service sample submission workflow
October 5, 2015Mass Spectrometry website: http://www.chem.ox.ac.uk/spectroscopy/mass-spec/ Page 14
White excel spreadsheet:‘Alternative ionisation service’
Yellow spreadsheet: ‘Accurate Mass service’1. 2.
1. Obtain an MSS or ESI number from sample login computer in ground floor open access MS room
2. Complete white sample submission form including your new MSS or ESI number (the form can be found under MS services on the MS website)
3. Label sample vial with MSS number or ESI number
4. Place labelled samples in tray in sample submission fridge
4. 3.
All results will be emailed to the email address provided on the submission form. Forms filled out incorrectly will not be processed.
http://www.chem.ox.ac.uk/spectroscopy/mass-spec/Services_allFiles/Services_sample%20submission%20forms.html
Proteomics service sample submission workflow
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2.If you have not used the proteomics service before you must first see Elisabete Pires to discuss your proteomics requirements.
Results will take up to 48 hours. You should contact Elisabete to check your analysis is completed successfully.
4.3.
Complete proteomics submission form
http://www.chem.ox.ac.uk/spectroscopy/mass-spec/Services_allFiles/Services_sample%20submission%20forms.html
1.
Bring sample and complete form down to basement MS room and submit to Elisabete in person. There is a fridge for storage of Samples.
Project and grant codes‘P2’ for part II students (unless you have a grant code)
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3. Trained user MS systems
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A range of trained-user MS systems are available for use once training in completed.
• Oligonucleotide MS• GC/MS• MALDI-MS• IC/MS• Native state-MS• Glovebox-MS• Quadrupole-MS• LC/MS• GC• Inorganic-MS
LC/MS
October 5, 2015Mass Spectrometry website: http://www.chem.ox.ac.uk/spectroscopy/mass-spec/ Page 18
2x LC/triple quadrupole‐MS : Useful for low resolution, high sensitivity and quantitation studies (basement & first floor)
LC‐TOF: Basement: Useful for separation of organic and inorganic small and large molecules
LC‐TOF: Useful for separation of organic and inorganic small and large molecules (1st floor)
LC‐TOF: Basement: Useful for separation and identification of Oligonucleotides
SYNAPT ion mobility MS:
• Complex mixture analysis• Amino acid analysis• Natural product identification• Metabolomics• Proteomics• Oligonucleotide MS
EP
Slide 18
EP5 Used to analyse/identified protein mixtures, modified peptides using nanoLC chromatography.Elisabete Pires, 22/09/2015
GC/MS
October 5, 2015Mass Spectrometry website: http://www.chem.ox.ac.uk/spectroscopy/mass-spec/ Page 19
GC/MS: Useful for high resolution, high sensitivity separation of compounds in the gas-phase with identification based on IE mass spectral libraries.
http://www.chem.ox.ac.uk/spectroscopy/mass-spec/Training_allFiles/Training_Intruments.html
Agilent GC/MS
• Complex mixture analysis• volatiles• SPME• Headspace analysis• Metabolomics
Native-state MS
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Q-TOF: Equipped with nano-ESI source robot (AdvionNanomate). Useful for analysis of native state proteins (with tertiary structure intact) and protein complexes.
http://www.chem.ox.ac.uk/spectroscopy/mass-spec/Training_allFiles/Training_Intruments.html
Waters Q‐TOF Micro
• Native state protein ID• PTM analysis• Binding assays
MALDI-TOF and MALDI-TOF/TOF instrument : Basement
October 5, 2015Mass Spectrometry website: http://www.chem.ox.ac.uk/spectroscopy/mass-spec/ Page 21
Waters MALDI micro
Matrix-Assisted Laser-Desorption Ionisation (MALDI) isa laser based ionisationtechnique useful for identificationof:
•Peptides/proteins•Oligonucleotides•Polymers. •Small molecules above 450 Da.
AB Sciex MALDI TOF-TOF
Trained user instrument access
October 5, 2015Mass Spectrometry website: http://www.chem.ox.ac.uk/spectroscopy/mass-spec/ Page 22
1. Discuss and agree your needs for MS with your supervisor.
2. Email or come and see James McCullagh or James Wickens who will arrange the training (~2 hours).
3. This will enable you to book and use the instrument unsupervised.
4. You may not use any trained user instruments unless you have received training from a member of the MS staff.
4.OtherExperiments
Orbitrap Exactive Plus
Stable isotope ratio mass spectrometry
Inert atmosphere MS
SYNAPT ion mobility MS
Probe MS Orbitrap Q‐Exactive
Mass Spectrometry website: http://www.chem.ox.ac.uk/spectroscopy/mass-spec/
Mass Spectrometry website: http://www.chem.ox.ac.uk/spectroscopy/mass-spec/
Prof James McCullaghHead of Research facility
Dr James WickensMS Services Manager
Mr Colin sparrowResearch Technician
Ms Elisabete PiresResearch Associate
Dr Ashley ShepherdResearch Associate
Areas of focusGCGC/MSMS of SolidsMALDIInorganic analysis
Areas of focusProteomicsMetabolomicsBiological MSLC/MSMALDISample prep
Areas of focusGCGC/MSMALDIAccurate Mass Analysis
Areas of focusGC/MSLC/MSOpen accessProbe MSOrbitrap MSSample prep
Areas of focusLC/MSMetabolomicsIsotope tracersIRMSIC-MSOrbitrap MSSample prep
Areas of specialism within the MS group
Data and its storage: MassStore
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MS experiments, particularly LC/MS and GC/MS create large data files. A typical proteomics data file for example can be > 200GB per sample.
Mass Spectrometry website: http://www.chem.ox.ac.uk/spectroscopy/mass-spec/
\\FILES-SUPPORT\Mass Spectrometry\MassStore-Research Groups
MS and NMR Software: MestReNova
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Mestrenova software is available for download from the chemistry server. This can be used to interrogate MS and NMR data, transform and collate for presentations. See MS website for further details about how to download.
Mass Spectrometry website: http://www.chem.ox.ac.uk/spectroscopy/mass-spec/
• Accurate Mass analysis• Identify LC/MS peaks• Interrogate open access results• Collate NMR and MS data
together• Annotate spectra• Produce reports
Protocol and safety in the MS labs
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1. No lab coats to be worn in any of the MS labs.
3. Open access data is backed up and stored for a period of 5 years. If you need
access to these data files please see a member of MS staff.
2. Never remove or modify any part of the mass spectrometer system, sample inlet
system or software. If something is not working properly please report to a member
of the MS facility staff.
4. Please read safety information on the mass spectrometry website and all
information provided at Mass Spectrometer terminals.
Mass spectrometry facility website
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Sign up to open access training
October 5, 2015Mass Spectrometry website: http://www.chem.ox.ac.uk/spectroscopy/mass-spec/ Page 29
Open access Training (Organic NMR and Mass Spectrometry):Sign up sheets at end of NMR talkMonday – Wed next week
Open Access Training for Inorganic users Tue – Wed sign up sheet other side of desk
Large molecule/protein open access training Please contact Dr James Wickens in the first instance (MS services Manager) to arrange training. [email protected]
Graduate Lecture Course in Mass Spectrometry (see website for schedule (8 lectures starting October 13th 2015).