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Servicios e Instalaciones Principales
Mass Spectrometry & Proteomics
Imagen
Core Facility Manager
Telephone:
+34 93 40 34869

Research information

Background

Modern proteomic tools are essential to understand biological questions. The Mass Spectrometry & Proteomics Core Facility (MSPCF) provides IRB Barcelona researchers and external users with a number of mass spectrometry-(MS) and proteomics-based techniques that can help to further their knowledge of disease, pathways, targets, and drug effects.

Set up in September 2007, the MSPCF Facility develops and implements a broad range of mass spectrometry-based methodologies and applies them to diverse topics in structural biology, biochemistry, cell biology and, into the clinics, in collaboration with IRB research groups and externals researchers. The Facility currently offers state-of-the-art applications in Quantitative Proteomics (differential expression protein analysis through label free, multiplexing and targeted strategies), High-throughput post-translational modifications (PTMs) analysis, Protein Interactions, Structural MS ((Biotinylation-MS, crosslinking-MS, Native MS; studying protein structure, protein complexes and their dynamic interactions) and Top-down High-throughput Proteomics (characterizing intact proteins, namely proteoforms).

The Facility provides consultancy services that cover the requirements of any scientific goal that calls for the use of MS analysis, from experimental design, experimental workflows, data acquisition and data analysis to result validation and interpretation. Feedback from users is considered crucial input for the continuous improvement of the Facility and its services. Samples are analysed either directly by Facility staff or by researchers (previously trained by facility members), who can use mass spectrometers through an open-access system.

The Facility team is composed by highly qualified researchers specialized in mass spectrometry techniques (from the sample preparation to the final data analysis and statistical treatment) and understands the service provided as being dynamic and requiring the continuous development and implementation of state-of-the-art methods and technologies in response to user needs. To reach these objectives, it pursues method development and subsequent implementation.

Equipped with high accuracy and high resolution mass spectrometers, the Facility has recently installed a new Orbitrap Eclipse which works attached to an Evosep One chromatograph. This instrumentation, together with the Orbitrap Fusion Lumos installed in 2016, allows challenging proteomics applications such as the analysis of low level PTMs, quantitative approaches using isobaric tags or metabolic labeling, as well as intact protein analysis and top-down characterization and a higher throughput capacity of analysis.

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The Facility has extensive expertise in proteomic analysis in all their approaches (e.g. intact proteins, classical proteomics, data interpretation and large data set analysis) and provides proteomics knowhow to the scientific community. Non-covalent protein-protein and protein-ligand interactions are also studied by MS and Ion mobility coupled to MS (IM/MS) in order to gain further insight into the mechanisms of action of these proteins in biological processes.

The Facilty currently leads the set-up of the Biomedical Proteomics Platform (IRB Barcelona’s Unit) for biomedical and translational Research usage in Catalonia, which is co-financed by the European Regional Development Fund (ERDF) in a join collaborative project with the Josep Carreras Leukemia Research Institute (FIJC), the Sant Joan de  Déu Foundation  (FSJD), the Vall Hebron Institute of Oncology (VHIO) and the University of Barcelona (UB) in the framework of the 2014-2020 ERDF Operational Progamme in Catalonia (Reference: IU16-015983).

The specialization and achievements of the Facility with respect to the implementation and development of Quantitative MS and Structural Proteomics workfows within the Spanish Proteomics community has led to its partnership in the PRBB network (Plataforma en red de Recursos Bioinformaticos y Biomoleculares, former ProteoRed), obtaining financial support from the Spanish Government-Instituto de Salud Carlos III for this purpose in the periods 2014-2017 and 2018-2020 (PRB3 (IPT17/0019 - ISCIII-SGEFI / ERDF), as a member group of Proteored, PRB3-ISCIII).  The Facility participates in the international Chromosome-Centric Human Proteome Project through the Spanish HPP consortium (SpHPP), In this regard, the Facility is contributing Top-down data to the “Chromosome 16 study”,

During the period 2014-2018, the IRB Barcelona Facility was granted a COST Action (European Cooperation in the field of Scientific and Technical Research-COST) from the Horizon 2020 program in a collaborative consortium initially formed by 8 European countries. The action was entitled “Native Mass Spectrometry and Related Methods for Structural Biology” (Action BM1403), aiming to bring together a group of researchers with a common interest, namely the development and application of new biomolecular MS methods in order to make the characterization of protein structure and dynamics more rapid and routine. The achievements of the Facility with respect to the implementation of structural MS approaches have positioned it as one of the pioneering labs undertaking this type of analysis in Spain. 

Please also see the selected publications section where the scientific contributions of the Facility are listed.

For more information, please contact the facility manager at marta.vilasecaarrobairbbarcelona.org

Selected publications

Gay M; Sánchez-Jiménez E; Villarreal L; Vilanova M; Huguet R; Arauz-Garofalo G; Díaz-Lobo M; López-Ferrer D; Vilaseca M
Methods In Molecular Biology
2044
( )
193 -
219
(2019)
Smith LM; Thomas PM; Shortreed MR; Schaffer LV; Fellers RT; LeDuc RD; Tucholski T; Ge Y; Agar JN; Anderson LC; Chamot-Rooke J; Gault J; Loo JA; Paša-Toli? L; Robinson CV; Schlüter H; Tsybin YO; Vilaseca M; Vizcaíno JA; Danis PO; Kelleher NL
Nature Methods
16
(10 )
939 -
940
(2019)
Molnar C; Heinen JP; Reina J; Llamazares S; Palumbo E; Breschi A; Gay M; Villarreal L; Vilaseca M; Pollarolo G; Gonzalez C
Science Advances
5
(8 )
eaaw7965 -
(2019)
Iacobucci C; Piotrowski C; Aebersold R; Amaral BC; Andrews P; Bernfur K; Borchers C; Brodie NI; Bruce JE; Cao Y; Chaignepain S; Chavez JD; Claverol S; Cox J; Davis T; Degliesposti G; Dong MQ; Edinger N; Emanuelsson C; Gay M; Götze M; Gomes-Neto F; Gozzo FC; Gutierrez C; Haupt C; Heck AJR; Herzog F; Huang L; Hoopmann MR; Kalisman N; Klykov O; Kuka?ka Z; Liu F; Maccoss MJ; Mechtler K; Mesika R; Moritz RL; Nagaraj N; Nesati V; Neves-Ferreira AGC; Iacobucci C; Piotrowski C; Aebersold R; Amaral BC; Andrews P; Bernfur K; Borchers C; Brodie NI; Bruce JE; Cao Y; Chaignepain S; Chavez JD; Claverol S; Cox J; Davis T; Degliesposti G; Dong MQ; Edinger N; Emanuelsson C; Gay M; Götze M; Gomes-Neto F; Gozzo FC; Gutierrez C; Haupt C; Heck AJR; Herzog F; Huang L; Hoopmann MR; Kalisman N; Klykov O; Kuka?ka Z; Liu F; Maccoss MJ
Analytical Chemistry
91
(11 )
6953 -
6961
(2019)
Vazquez de la Torre, Aurelio; Gay, Marina; Vilaprinyo-Pascual, Silvia; Mazzucato, Roberta; Serra-Batiste, Montserrat; Vilaseca, Marta; Carulla, Natalia
Analytical Chemistry
90
(7 )
4552 -
4560
(2018)
Nadal M; Prekovic S; Gallastegui N; Helsen C; Abella M; Zielinska K; Gay M; Vilaseca M; Taulès M; Houtsmuller A; Van Royen M; Claessens F; Fuentes-Prior P; Estébanez-Perpiñá E
Nature Communications
8
(14388 )
14388 -
(2017)
Izquierdo-Serra M; Bautista-Barrufet A; Trapero A; Garrido-Charles A; Diaz-Tahoces A; Camarero N; Pittolo S; Valbuena S; Perez-Jimenez A; Gay M; Garcia-Moll A; Rodriguez-Escrich C; Lerma J; De La Villa P; Fernandez E; Pericas M; Llebaria A; Gorostiza P
Nature Communications
7
(12221 )
12221 -
(2016)
Marin-Montesinos I; Paniagua J; Vilaseca M; Urtizberea A; Luis F; Feliz M; Lin F; Van Doorslaer S; Pons M
Physical Chemistry Chemical Physics
17
(8 )
5785 -
5794
(2015)
Pujol-Pina R; Vilaprinyó-Pascual S; Mazzucato R; Arcella A; Vilaseca M; Orozco M; Carulla N
Scientific Reports
5
(14809 )
14809 -
(2015)
Vilà-Rico M; Colomé-Calls N; Martín-Castel L; Gay M; Azorín S; Vilaseca M; Planas A; Canals F
Journal Of Proteomics
127
(pt b )
234 -
246
(2015)
Slamnoiu S; Vlad C; Stumbaum M; Moise A; Lindner K; Engel N; Vilanova M; Diaz M; Karreman C; Leist M; Ciossek T; Hengerer B; Vilaseca M; Przybylski M
Journal Of The American Society For Mass Spectrometry
25
(8 )
1472 -
1481
(2014)
Branowska D; Chaciak B; Siuchta O; Olender E; Ledwon P; Lapkowski M; Poronik E; Wysocki W; Karczmarzyk Z; Skorka L; Filapek M; Krompiec S; Urbanczyk-Lipkowska Z; Kalicki P; Aresu E; Fioravanti S; Pellacani L; Sciubba F; Trulli L; López A; Tarragó T; Vilaseca M; Giralt E; Saßmannshausen J; Klett J; Kennedy AR; Parkinson JA; Armstrong D
New Journal Of Chemistry
37
(12 )
4250 -
(2013)
Amata I; Maffei M; Igea A; Gay M; Vilaseca M; Nebreda A; Pons M
Chembiochem
14
(14 )
1820 -
1827
(2013)
López A; Tarragó T; Vilaseca M; Giralt E
New Journal Of Chemistry
37
(5 )
1283 -
1289
(2013)
Borges-Alvarez M; Benavente F; Vilaseca M; Barbosa J; Sanz-Nebot V
Journal Of Mass Spectrometry
48
(1 )
60 -
67
(2013)
Arcella A; Portella G; Ruiz M; Eritja R; Vilaseca M; Gabelica V; Orozco M
Journal Of The American Chemical Society
134
(15 )
6596 -
6606
(2012)
Bonet-Costa, Carles; Vilaseca, Marta; Diema, Claudio; Vujatovic, Olivera; Vaquero, Alejandro; Omenaca, Nuria; Castejon, Lucia; Bernues, Jordi; Giralt, Ernest; Azorin, Fernando
Journal Of Proteomics
75
(13 )
4124 -
4138
(2012)
Flores-Morales P; Diema C; Vilaseca M; Estelrich J; Luque F; Gutiérrez-Oliva S; Toro-Labbé A; Silva E
Bioorganic & Medicinal Chemistry
19
(5 )
1613 -
1622
(2011)
Sánchez L; Madurga S; Pukala T; Vilaseca M; López-Iglesias C; Robinson C; Giralt E; Carulla N
Journal Of The American Chemical Society
133
(17 )
6505 -
6508
(2011)
Casado-Vela J; Cebrián A; Gómez Del Pulgar M; Sánchez-López E; Vilaseca M; Menchén L; Diema C; Sellés-Marchart S; Martínez-Esteso M; Yubero N; Bru-Martínez R; Lacal J
Proteomics
11
(4 )
590 -
603
(2011)
Borg J; Campos A; Diema C; Omeñaca N; De Oliveira E; Guinovart J; Vilaseca M
Clinical Proteomics
8
(1 )
6 -
(2011)
Ferrer M, Pedrosa A, Rodríguez L, Rossell O, Vilaseca M
Inorganic Chemistry
49
(20 )
9438 -
49
(2010)

Projects

Plataforma de apoyo a la Investigación en Ciencias y Tecnología de la salud de la convocatoria de 2017 de la Acción Estratégica en Salud 2013-2016, del Instituto de Salud Carlos III (ISCIII), pudiendo estar cofinanciada con cargo al Fondo Europeo de Desarrollo Regional (FEDER). Referencia: PT17/0019/0022 (Plataforma de Proteomica, Genotipado y Líneas celulares, PRB3)

Gobierno de españa
European union
Instituo de Salud Carlos III
PR3

Plataforma de apoyo a la Investigación en Ciencias y Tecnología de la Salud, cofinanciada por la Acción Estratégica en Salud 2013-2016, Instituto de Salud Carlos III, y pudiendo estar cofinanciada con cargo al Fondo Europeo de Desarrollo Regional (FEDER). Referencia: PT13/0001/0015 (Plataforma de Recursos Biomoleculares y Bioinformáticos, PRB2)

Gobierno de españa
European union
Instituo de Salud Carlos III

Cost Action “Native Mass Spectrometry and Related Methods for Structural Biology”, financiada por COST (European Cooperation in Science and Technology). Referencia: BM1403

COST
Horizon 2020

Set up and development of the Biomedical Proteomics Platform (IRB Barcelona’s Unit) for biomedical and translational Research. Co-financed by the European Regional Development Fund (ERDF), in the framework of the 2014-2020 ERDF Operational Progamme in Catalonia, with €519,500. Reference: IU16-015983.

Secretaria d'Universitas i Recerca

Identificació i validació de biomarcadors inicials en la fase aguda de la COVID-19 Predictors d'evolució” (IVETTE) con el apoyo de la Fundació La Marató de TV3 (202113-30-31-32).

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Services

The services offered include MS, MS/MS and MSn analysis using atmospheric pressure ionisation techniques (electrospray and nanoelectrospray) coupled to LC, nanoLC or infusion inlets.

 

The Facility has the following lines of expertise:

  • Protein and Peptide Analysis
    • Identification/characterisation
      • Intact protein analysis
      • High-throughput proteomics, PTM analysis
        • Bottom-up proteomics
        • Top-down proteomics, Middle-down proteomics
      • mAbs analysis (ADC)

    • Quantitative Proteomics
      • Differential protein expression analysis
        • Label-free strategies (DDA, DIA)
        • Multiplexing strategies (iTRAQ, TMT SPS MS3, SILAC)
      • Targeted analysis (PRM)

    • Structural Proteomics
      • Native MS, IM-MS, Non covalent interactions
      • HDX-MS, Cross-linking MS, Limited proteolysis MS
      • Affinity MS (SAW-MS)

  • Small Molecule Analysis
    • Characterisation (on final products, exact mass and MS/MS)

 

Equipment and specialised applications

Orbitrap Eclipse (Thermo Scientific) (NEW 2021) 

Orbitrap Eclipse (Thermo Scientific)

The capacity of the Facility was significantly strengthened by the acquisition of a new generation tribrid mass spectrometer that includes a quadrupole, a linear ion trap, and an Orbitrap mass analyzer. This instrument allows multiple fragmentation techniques—CID, HCD, and ETD—during any stage of MSn, with subsequent mass analysis in either the ion trap or Orbitrap mass analyser.

The instrument takes advantage of the combination of a brighter ion source than previous Orbitrap instruments, with a higher capacity transfer tube and an electrodynamic ion funnel for increased sensitivity. It is provided with an advanced active ion beam guide for improved robustness, a segmented quadrupole, and improved ion transmission and ETD performance.

Advanced vacuum technology allows tuning of the vacuum in the IRM cell to improve ion transmission and S/N for intact proteins, as well as for their fragmentation.

This instrument is used in the most challenging applications in proteomics, both in Top-down and Bottom-up approaches. It is used coupled to two nanoLC systems depending on the MS approach: a Dionex Ultimate 3000 nanoLC sytem (Thermo) or a NanoAcquity UPLC from Waters.  A chip-based nanoESI interface (Advion Triversa Nanomate) is used in both cases.

 

Evosep One (New 2021)

 

Evosep One

The chromatograph Evosep One, which we use coupled to the Orbitrap Eclipse, is designed to be able to work with fast chromatographies and therefore provides real  high-throughput and robust analysis, being ideal for clinical proteomic applications.
This nanoLC uses Evotips as disposable trap columns, where samples are loaded and desalted offline. The autosampler picks up the tip and integrates elution with liquid chromatography. This removes sample handling steps and reduces injection cycle overheads. Minimal wear and tear on mechanical components is ensured as all elution and gradient formation happen at low pressure. The Evotip is left for disposal after each run, which then allows partial elution to extend column lifetime. Analytically important peptides are effectively retained, and cross-contamination reduced.
 
The Evosep sytem facilitates the LC-MS/MS analysis of large number of samples to be able to undertake efficiently the analysis of sample cohorts in clinical and translational applications.

 

Orbitrap Fusion Lumos (Thermo Scientific) (2016) 

 

The capacity of the Facility was significantly strengthened by the acquisition in 2016 of this tribrid mass spectrometer that includes a quadrupole, a linear ion trap, and an Orbitrap mass analyzer. This instrument allows multiple fragmentation techniques—CID, HCD, and ETD—during any stage of MSn, with subsequent mass analysis in either the ion trap or Orbitrap mass analyser.
The instrument takes advantage of the combination of a brighter ion source than previous Orbitrap instruments, with a higher capacity transfer tube and an electrodynamic ion funnel for increased sensitivity. It is provided with an advanced active ion beam guide for improved robustness, a segmented quadrupole, and improved ion transmission and ETD performance.
Advanced vacuum technology allows tuning of the vacuum in the IRM cell to improve ion transmission and S/N for intact proteins, as well as for their fragmentation.
This instrument is used in the most challenging applications in proteomics, both in Top-down and Bottom-up approaches. It is used coupled to two nanoLC systems depending on the MS approach: a Dionex Ultimate 3000 nanoLC sytem (Thermo) or a NanoAcquity UPLC from Waters.  A chip-based nanoESI interface (Advion Triversa Nanomate) is used in both cases.

 

 

Synapt High Definition MS System (Waters-Micromass)

Synapt High Definition MS System (Waters-Micromass) (2007). Hybrid QTOF instrument with an incorporated Triwave Cell. This instrument allows tandem MS to be combined with ion mobility, thus permitting the analysis of samples differentiated not only by their mass to charge ratio, but also by their shape and size. The instrument is used to analyse the macromolecular structure and conformation of intact proteins and to study non-covalent interactions. When working in its normal configuration, the instrument is attached to a chip-based nanoESI interface (Advion Triversa Nanomate).

 

 

Premier XE

LCT-Premier XE (Waters-Micromass) (2007). Orthogonal acceleration time-of-fight mass spectrometer ideal for the analysis of high molecular weight compounds. This instrument has been modified to achieve inert conditions inside the ionization source, in order to allow amide H/D exchange experiments for the study of the dynamic and structural properties of proteins and their complexes. In the standard set-up configuration it works coupled to an LC device (UPLC Acquity, Waters), e.g large protein analysis (mAbs).-

 

Complementary devices

The Facility is equipped with the following complementary devices, in addition the chromatographic systems already mentioned:

 

A Gelfree (Gel-Eluted Liquid Fraction Entrapment Electrophoresis) system for protein separation:

Gelfree 8100 Fractionation system (Protein Discovery Inc.) (2010)

 

More info

Before sample submission

Contact the MS team. We advise you on the best analytical methodology for your specific application.

 

Instructions for sample delivery

One of the most important steps to achieve a successful result in mass spectrometry analysis is a well-planned experiment involving an adequate sample preparation for each MS analysis. Specially, for proteomic experiments is strongly recommended to contact with MS team before starting it. Here you have some general instructions, but do not hesitate to contact us for any doubt.

 

General features

  1. Work in a clean environment.
  2. Avoid keratin contamination by using gloves and lab coat.
  3. Do not use autoclaved material.
  4. Wash all the material with milliQ water and MeOH (high quality) and let them dry before use.
  5. Use LC-MS grade reagents.
  6. If possible use low-binding eppendorf.
  7. Use simple and well-recognized tube labeling. We recommend using consecutive numbers.

 

Samples for in-gel digestion

  1. When possible run a 0.75 mm thickness gel.
  2. Use clean recipients for staining. Close them during staining period to avoid contamination.
  3. NEVER use recipients that were previously used for Western blot.
  4. Use a mass spectrometry compatible protocol for gel staining.
  5. If you use fluorescent stain do not cut the bands manually by using a transilluminator.
  6. Cut only the stained region. The higher protein concentration and the lower acrylamide presence, the better protein identification.
  7. When possible cut the gel spots or bands in a laminar flow hood to avoid keratin contamination.
  8. Store bands at 4ºC without water or other solvents.

 

Samples for in-liquid digestion

  1. There are some reagents not compatible with mass spectrometry. Below are listed the most commonly used and not compatible with LC-MS. Please tell us the reagents you use for sample preparation.
    • Igpal/NP40 is almost impossible to remove and shows large polymers along all the chromatogram, so we will suggest changing it for N-octyl-β-glucopyranoside or avoiding any detergent if possible.
    • The RNAse is a protein that could mask the proteins of interest.
    • When possible avoid glycerol.
  2. For IP experiments a negative and a positive control are needed to find out nonspecific interactions. We also recommend running a SDS-PAGE gel with 10% of the sample to have an idea how the IP works.
  3. The amount of sample required for protein identification depends on sample complexity and purity. For complex samples at least 1nM is required.

 

Intact proteins

  1. An efficient ionization (ESI or nanoESI) of proteins depends on their length, and physicochemical properties (pKa, solubility, structure and hydrophobicity). It is possible to work under denaturing or under non-denaturing conditions
    • Denaturing conditions: proteins are solubilized generally in a 1:1 mixture of acetonitrile/water or methanol/water and 1% formic acid is added to enhance ionization (all solvents must be MS quality). Depending on their purity, proteins are analyzed by infusion or by LC-MS. 1μM to 10 μM protein concentration is required (volume: 10-100 μl). Low dynamic ranges are normally achieved and pre-purification is therefore required. Non-volatile salt removal is necessary (Zip-tips can be used). Maximum non-volatile salt accepted in the case of LC-MS experiments is 150 mM. Infusion experiments require complete salt removal.
    • Non-denaturing conditions: proteins or non-covalent protein complexes are solubilized in NH4OAC (10mM-1M). Salt removal is imperative and is done normally by dialysis or with Biospin columns. 5μM to 50 μM protein concentration is required.
  2. Top-down (MS/MS experiments) can be performed on intact proteins to localize modifications or confirm amino acidic sequence. Protein length is limited to 30 kDa to perform MS/MS in the gas phase with the aim of entirely sequencing the protein. Middle-down strategies are otherwise used. Denaturing conditions are used in this case. 5μM to 10 μM protein concentration is required (100 μl).
  3. Top-down can also be used for protein identification (up to 100-150 kDa). In this case protein coverage is low (this strategy is very much in underdevelopment with respect to the classical bottom-up approach (proteomics) although useful for protein isoform identification )

 

Sample submission/reception

There are two ways to submit samples for MS analysis:

  1. Submission to the technician-run service by filling in the sample/s request form (available here or in the MSC lab).
  2. Self-service (open-access) systems. Only for researchers qualified and trained to operate the mass spectrometers.
    If you plan to use the MS service often and would like to be considered for training, please contact Dr. Marta Vilaseca.
 

Custody and maintenance of samples

When filling the request form, your sample or set of samples will be identified by a code number (apart from the users label). Depending on the analysis requested it will be classified in the corresponding "to-run" folder for each instrument.

The sample will be also classified and kept in the conditions specified in the request form.

If preferred you can keep your sample and bring it a day before the beginning of analysis (following scheduled dates).

 

Time of response for an analysis

There is a calendar (contact the facility manager) for each instrument and submissions will be ordered by time of entry (IRB Barcelona groups are given preferential treatment).

Given the diversity of applications, some of which require only fast analysis while others require longer analysis time, the calendar will be organised as follows:

  • Users can reserve a week in a specific instrument for long application (maximum 2 full weeks per user, depending on demand).
  • Between long time applications we always reserve a week to perform shorter analysis in order to avoid delays.
  • Mondays (or a pre-specified day) are reserved for the maintenance of the equipment and set up of the required technique pretended to use that week in each instrument.
 

Results delivery

Results performed by MS staff will be given in printed format (spectra and report specifying analytical conditions). Digital format data can also be kept if requested.

If further processing of your results is required a computer will be reserved in the facility for this purpose.

Our future goal is that users can also access data via a secured network. Data will be transferred to a common-server and back-ups will be performed automatically by the ITS Department. Data will be kept for a maximum of two years.

 

Contact


Mass Spectrometry Request Form

e-mail: marta.vilasecairbbarcelona.orgmarina.gayirbbarcelona.org

Telephone: +34 934039815

Group members

Core Facility Manager
Senior Research Officer
Research Officer
Research Officer
Research Officer
Research Officer
Research Officer

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