
Core facilities
PROMIFA

Mission
The Protein Microsequencing Facility provides different analytical services aimed at structurally characterizing small molecules, peptides and proteins. The Facility is equipped with state-of-the-art mass spectrometers. Access to mass spectrometry is a crucial key in the emerging field of proteomics and metabolomics, which are giving a major contribution to the description and understanding of biological processes at the molecular level.
Services
The goal of the Facility is to assist scientists in their research on a fee-for-service basis, covering all the operating costs, including consumables, time-machine and technical support.
PROMIFA services can be divided in:
- Biochemical Services
- Mass Spectrometry Services
- Proteomics Services
- Metabolomics Services
All the services comprise appropriate scientific consultancy, interpretation of the results, a report on the analytical data signed by the service coordinator (on demand).
Guidelines and detailed protocols for sample preparation are available here
Request form for internal and external users are available here:
Access info
Due to the complexity and cost of some of the equipment, only the staff of the facility has direct access to the machines. Interested people can be trained upon request. Samples are accepted every day.
PROMIFA Facility is located in Dibit 2, C1, 4Th floor.
First time users and anyone wanting to submit large numbers of samples at once (more than 15) are urged to contact the Facility prior to sample submission. The sample(s) has to be shipped together with the filled request form/order. PROMIFA strongly suggest contacting the staff of the Facility before filling in any request form, in order to define exactly the activity to be carried out on the samples.
Here you can find some frequently asked questions: FAQs.
Protein characterization:
- protein/peptide analysis by HPLC/MS and MS/MS;
- sample purification (zip tip or similar) and intact protein MW determination by nanospray off line MS.
Protein identification: PMF and sequencing
- PTM-characterization: validation by Western Blotting and 2DE +/- removal of the PTM (glycosylation, phosphorylation);
- phosphorylation characterization by TiO2 or IMAC enrichment and mass spectrometry;
- identification of a spot or band by in gel digestion and LC-MS/MS .
Quantitative protein profiling by mass spectrometry and statistical analysis. Differential protein display in two or more diverse conditions using:
- iTRAQ or TMT labelling of peptides,
- SILAC methodology,
- Label-free approach.
Biomarker profiling in biological samples by 2DE, image analysis and statistical evaluation. Differential protein display in two or more diverse conditions in plasma, serum, urine, tissue.
Metabolomics/Lipidomics by mass spectrometry and statistical analysis. Untargeted workflow for differential small molecules/lipids display in: plasma, serum, urine, tissues.
Quantitative Metabolomics/Lipidomics by mass spectrometry and statistical analysis. Targeted workflow for specific small molecules/lipids display in plasma, serum, urine, tissues.
Protein analysis |
Internal/external |
Protein/peptide analysis by HPLC/MS and MS/MS | 100/110 € |
Sample purification (zip tip or similar) and intact protein MW determination by nanospray off line MS |
75/82 € |
Protein identification: PMF and sequencing | |
PTM-characterization: validation by Western Blotting and 2DE +/- removal of the PTM (glycosylation, phosphorylation) | 500/600 € |
Phosphorylation characterization by TiO2 or IMAC enrichment and mass spectrometry | 300/350 € |
Identification of a spot or band by in gel digestion and LC-MS/MS | 200/240 € (the price goes down with the number of bands/spots) |
Biomarker profiling in biological samples by 2DE, image analysis and statistical evaluation | |
Differential protein display in control vs patient biological samples: plasma, serum, urine, tissue. Six samples: control vs treated/patient in triplicates |
1500/1800 € (800/960€ for each additional condition) |
Quantitative protein profiling by mass spectrometry and statistical analysis | |
iTRAQ or TMT labelling of peptides (run in technical duplicate) | 2500/3000€ |
SILAC methodology (run in technical duplicate) | 2500/3000€ |
Label-free approach (run in technical triplicate) (iTRAQ or TMT-reagents or labelled amino acids are in charge to the customers) |
800/960€ |
Metabolomics/Lipidomics by mass spectrometry and statistical analysis | |
Untargeted workflow for differential small molecules/lipids display in: plasma, serum, urine, tissues (C18, HILIC, C8, positive and negative polarity). | Prices are tailored to each project |
Quantitative Metabolomics/Lipidomics by mass spectrometry and statistical analysis | |
Targeted workflow for specific small molecules/lipids display in: plasma, serum, urine, tissues. | Prices are tailored to each project |
Access to MS | 15/25€/h |
Data analysis | 60/80€/h |
ACTIVITIES |
PROCESSING TIME |
Protein characterization | |
Protein and peptide analysis by HPLC/MS and MS/MS | 3 days |
Sample purification (zip tip or similar) and intact protein MW determination by MALDI or nanospray off line MS | 3 days |
Protein identification: PMF and sequencing | |
PTM-characterization: validation by Western Blotting and 2DE +/- removal of the PTM (glycosylation, phosphorylation) | 2 weeks |
Phosphorylation characterization by TiO2 or IMAC enrichment and mass spectrometry | 2 weeks |
Identification of a spot or band by in gel digestion and LC-MS/MS |
1 week |
Processing timing depends on the number of samples for:
- Biomarker profiling in biological samples by 2DE, image analysis and statistical evaluation. Differential protein display in two or more diverse conditions in: plasma; serum; urine; tissues.
- Quantitative protein profiling by mass spectrometry and statistical analysis. Differential protein display in two or more diverse conditions using: iTRAQ or TMT labelling of peptide; SILAC methodology;Label-fre approach.
- Metabolomics/Lipidomics by mass spectrometry and statistical analysis. Untargeted workflow for differential small molecules/lipids display in: plasma; serum; urine; tissues.
- Quantitative Metabolomics/Lipidomics by mass spectrometry and statistical analysis. Targeted workflow for specific small molecules/lipids display in: plasma; serum; urine; tissues.
In case of maintenance or out of order instrumentation as well as when multiple samples are submitted, the processing time can be longer.
Output of the results will be sent in an Excel or Word format, along with information on: mass spectrometer settings, protein database version, search parameters, experimental conditions for 2DE, constraints for image analysis and statistical evaluation.
When the Facility will send the results of the analysis (by fax or e-mail) then the Grant Office will give the invoice to the user for the payment.
Ferrari A, Longo R, Fiorino E, Silva R, Mitro N, Cermenati G, Gilardi F, Desvergne B, Andolfo A, Magagnotti C, Caruso D, Fabiani E, Hiebert SW, Crestani M. HDAC3 is a molecular brake of the metabolic switch supporting white adipose tissue browning. Nat Commun. 2017 Jul 21;8(1):93. doi: 10.1038/s41467-017-00182-7.
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Barbariga M, Curnis F, Andolfo A, Zanardi A, Lazzaro M, Conti A, Magnani G, Volontè MA, Ferrari L, Comi G, Corti A, Alessio M. Ceruloplasmin functional changes in Parkinson’s disease-cerebrospinal fluid. Mol Neurodegener. 2015 Nov 4; 10: 59. doi: 10.1186/s13024-015-0055-2.
D’Addio F, La Rosa S, Maestroni A, Jung P, Orsenigo E, Ben Nasr M, Tezza S, Bassi R, Finzi G, Marando A, Vergani A, Frego R, Albarello L, Andolfo A, Manuguerra R, Viale E, Staudacher C, Corradi D, Batlle E, Breault D, Secchi A, Folli F, Fiorina P. Circulating IGF-I and IGFBP3 levels control human colonic stem cell function and are disrupted in diabetic enteropathy. Cell Stem Cell. 2015 Oct 1;17(4):486-98. doi: 10.1016/j.stem.2015.07.010.
Barbariga M, Curnis F, Spitaleri A, Andolfo A, Zucchelli C, Lazzaro M, Magnani G, Musco G, Corti A, Alessio M. Oxidation-induced structural changes of ceruloplasmin foster NGR motif deamidation that promotes integrin binding and signaling. Biol. Chem. 2014 Feb 7; 289(6):3736-48. doi: 10.1074/jbc.M113.520981. Epub 2013 Dec 23.
Magagnotti C, Matassa PG, Bachi A, Vendettuoli V, Fermo I, Colnaghi MR, Carletti RM, Mercadante D, Fattore E, Mosca F, Andolfo A. Calcium signaling-related proteins are associated with broncho-pulmonary dysplasia progression. Proteomics. 2013 Dec 6; 94: 401-12. doi: 10.1016/j.jprot.2013.10.007. Epub 2013 Oct 16.

Thermo Scientific LTQ Orbitrap XL mass spectrometer equipped with an Easy nLC Proxeon Biosystems

Thermo Scientific Q-Exactive mass spectrometer equipped with nUPLC Proxeon Biosystems

AB-Sciex TripleTOF5600+ mass spectrometer equipped with on-line UPLC Agilent 1290 and NanoMate Triversa (avion)