Graduate course in mass spectrometry starting 13th Oct ... · PDF fileGraduate course in mass spectrometry starting ... Graduate Course in Mass Spectrometry starting 13th Oct

October 6, 2016Page 1

Graduate course in mass spectrometry starting 13th Oct.

All part IIs, graduate students & postdocs welcome.

8 lectures in Michaelmas term, Wolfson seminar room 11am, Thursdays.

1. Oct 13th Introduction to mass spectrometry and basic principles2. Oct 20th Mass Analysers: Quadrupoles, Time of Flight, Orbitraps and hybrid instruments.3. Oct 27th Collisions in vacuum: cooling, activating, and sizing ions. 4. Nov 3rd Ion Chemistry in mass spectrometry 5. Nov 10th Separation methods (chromatography) coupled to mass spectrometry. 6. Nov 17th Proteomics break7. Dec 1st Metabolomics8. Dec 8th Mass spectrometry for biophysics and structural biology.

Graduate Course in Mass Spectrometry starting 13th Oct.

DEPARTMENT OF CHEMISTRYMASS SPECTROMETRY RESEARCH FACILITY

Professor James McCullagh, Head of the Mass Spectrometry Research Facility

INTRODUCTION TO THE CRL MASS SPECTROMETRY RESEARCH FACILITY

Introduction

Mass Spectrometry website: http://www.chem.ox.ac.uk/spectroscopy/mass-spec/

1. MS overview2. People3. Facilities4. Instrument locations5. Open access MS6. Trained users MS7. Project work8. Software and data storage9. Lectures and events

Identification Chemical formula prediction

260.0297185↓

C6H13O9P↓

Glucose-1-phosphate

What can mass spectrometry do for you?


Structural analysisFragmentation studies using MS

Quantification Relative or absolute

LC/MS, GC/MS

Different types of MSInlets, ionisation, sensitivity accuracy

People in the Mass Spectrometry Research Facility


Prof James McCullaghHead of Research facility

Dr James WickensMS Services Manager Research Associate

Ms Elisabete PiresResearch Associate

22 MS systems4 full time staffMultiple services

Dr Ashley ShepherdResearch Associate

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MS facility and staff support Research in the CRL (Part II, DPhil, Postdoc and PIs).

Location of main MS labs in CRL


Basement Mass Spec Lab 2

Basement Mass Spec Lab 1

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 Department 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.

Dr James Wickens Mass Spectrometry

Services Manager

Types of access to mass spectrometry


Dr James Wickens Mass Spectrometry

Services Manager

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

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

OPEN ACCESS LC/MS1. Fast column chromatography LC/MS2. <10mins per sample

Small molecule open access

Protein open access

Ground floor MS room

Ground floor MS room

1. Open access results


Small molecule open access Protein open access

Small moleculesResults are emailed as a PDFRaw data files available from3am next day on the chemistrynetwork: \\FILES-SUPPORT\Mass Spectrometry\Open Access

Protein Open AccessResults deposited as PDF in sepárate folder in same location.

Results: PDF emailed Results: PDF emailed

Adducts M+HM+NaM-H

1. Location of open access systems: ground floor CRL


All open access MS systems are located in the ground MS room

2. Small molecule characterisation services


SMALL MOLECULES (ORGANIC AND INORGANIC COMPOUNDS):1. Accurate Mass Service (novel compounds)2. High resolution Service (unable to obtain results by open access)

Small molecules sample login: Ground floor Lab 20.062 (same as open access)

Dr James Wickens Mass Spectrometry Services Manager

2. Sample submission


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

2.Large molecule characterisation services


BIOLOGICAL MASS SPECTROMETRY:1. Proteomics Services2. Native state protein MS3. Metabolite profiling

Biological MS services: Submit samples to Elisabete Pires in basement

[email protected]

2. Proteomics and biological MS service sample submission


2.If you have not used biological service before you must first see Elisabete Pires to discuss your proteomics requirements.

Results will take longer that for small molecule servces. You should contact Elisabete to check your analysis has completed successfully.

4.3.

Complete proteomics submission form

http://www.chem.ox.ac.uk/spectroscopy/mass-spec/Services_allFiles/Services_sample%20submission%20forms.html

[email protected]

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.

2.Project grant codes


‘P2’ for part II students (unless you have a grant code)

3. Trained user MS systems


A range of trained-user MS systems are available for use once training in completed.

• LC/MS• GC/MS• Oligonucleotide MS• MALDI-MS• IC/MS• Native state-MS• Glovebox-MS• Quadrupole-MS• GC• Inorganic-MS

3. Liquid chromatography coupled to MS


2x LC/triple quadrupole‐MS : Useful for low resolution, high sensitivity and quantitation studies (basement & first floor)

LC‐TOF: Useful for separation of organic and inorganic small and large molecules (1st floor)

SYNAPT ion mobility MS

• Complex mixture analysis• Amino acid analysis• Natural product identification• Metabolomics• Proteomics• Oligonucleotide MS

3. Gas chromatography coupled to MS


GC/MS: Useful for high resolution, high sensitivity separation of compounds in the gas-phase with identification based on IE mass spectral libraries.

Agilent GC/MS

• Complex mixture analysis• volatiles• SPME• Headspace analysis• Metabolomics

http://www.chem.ox.ac.uk/spectroscopy/mass-spec/Training_allFiles/Training_Intruments.html

Native‐state protein MS


Q-TOF: Equipped with nano-ESI source robot (AdvionNanomate). Useful for analysis of native state proteins (with tertiary structure intact) and protein complexes.

Waters Q‐TOF Micro

• Native state protein ID• PTM analysis• Binding assays

http://www.chem.ox.ac.uk/spectroscopy/mass-spec/Training_allFiles/Training_Intruments.html

3. MALDI‐TOF & MALDI TOF‐TOF


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

3. Trained user instrument access


1. Discuss and agree training is necessary with your supervisor.

2. Email or come and see James McCullagh or James Wickens who will arrange the training which takes 2-3 hours.

3. Once completed you will be able 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. Other instruments


4.OtherExperiments

Orbitrap Exactive Plus

Stable isotope ratio mass spectrometry

Inert atmosphere MS

SYNAPT ion mobility MS

Probe MS Orbitrap Q‐Exactive

Areas of focus within the MS facility team


Prof James McCullaghHead of Research facility

Dr James WickensMS Services Manager

New person Research Associate

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

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Mass Spectrometry Data Files: MassStore


MS experiments, particularly LC/MS and GC/MS create large data files. A typical proteomics data file for example can be > 200GB per sample.

\\FILES-SUPPORT\Mass Spectrometry\MassStore-Research Groups

MS and NMR Software: Mestrenova


Mestrenova software is available for download from the Mass Spectrometry and NMR websites. 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.

• Accurate Mass analysis• Identify LC/MS peaks• Interrogate open access results• Collate NMR and MS data

together• Annotate spectra• Produce reports

Protocols and Safety in the MS labs


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


Training courses and Lectures in MS


Open access Training (Organic NMR and Mass Spectrometry):Sign up sheets at end of NMR talkThursday 6th and Friday 7th

Open Access Training for Inorganic users Monday 10th sign up sheet other side of desk

Large molecule/protein open access training Please contact Elisabete Pires 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 2016).

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Graduate course in mass spectrometry starting 13th Oct ... · PDF fileGraduate course in mass spectrometry starting ... Graduate Course in Mass Spectrometry starting 13th Oct