Flow Cytometry Core
Maria Grazia Totaro
Maria Grazia obtained her Master's degree in Biology Applied to Biomedical Research from the University of Insubria in Varese. Her academic and professional training was enriched through experiences at prestigious research institutes such as the Institute for Research in Biomedicine (IRB) in Bellinzona and the European Institute of Oncology (IEO) and Humanitas Institute in Milan, where she collaborated with leading researchers in the field of immunology.
During her PhD at Humanitas Institute, her passion for immunology led her to explore the fascinating world of flow cytometry and its countless applications.
After earning her PhD, Maria Grazia applied her expertise by joining the Imaging Unit at IFOM, led by Dr. Dario Parazzoli, as a flow cytometry specialist. Over the course of 10 years, she honed her skills and significantly contributed to the growth and development of the unit's flow cytometry activities, becoming a key reference figure.
In October 2024, she was appointed Head of Facility of the Flow Cytometry Unit, established following the division of the Imaging Unit, taking on the responsibility of leading this new entity towards scientific excellence.
Gallery Flow Cytometry unit
Facility members
Michela Cravedi
Sara Martone
Reference PI:
Massimiliano Pagani
Unit info
Since its introduction in the 1960s, flow cytometry has become an essential tool in biomedical research, offering rapid data acquisition and in-depth analysis of multiple biological parameters. Recent technological advancements, particularly the introduction of full-spectrum flow cytometry, have significantly increased the number of parameters that can be analyzed simultaneously—now exceeding 50.
By enabling the precise identification and isolation of distinct cell populations, flow cytometry plays a crucial role in advancing cancer research. From uncovering the mechanisms of tumor progression to identifying potential therapeutic targets, this technology continues to drive discoveries that deepen our understanding of cancer biology and pave the way for innovative treatments.
At IFOM's Flow Cytometry Core Facility, we provide cutting-edge technology, state-of-the-art instrumentation, and expert support for a wide range of applications, including:
- Immunophenotyping
- Cell cycle and proliferation analysis
- Metabolic studies
- Apoptosis detection
- Fluorescent protein expression analysis
- Cytokine, chemokine, and intra/extracellular protein detection
Available Instrumentation
Researchers have access to both conventional and spectral flow cytometers:
- Traditional Flow Cytometers: FACSCanto II, Attune NxT
- Spectral Flow Cytometer: Aurora 5L
Independent instrument use is permitted after completing theoretical and practical training provided by the facility.
For cell sorting, access is available only with assistance on the following instruments:
- Conventional Cell Sorters: MoFlo Astrios, FACSAria IIU
- Spectral Cell Sorter: Aurora CS
Applications/Services
Services
- Instrument setup, optimization, and maintenance
- Assistance with experiment design, sample preparation, and gating strategy
- Access to cutting-edge technologies and reagents through industry collaborations
- Networking with other core facilities to stay at the forefront of innovation
- Data analysis using multiple software platforms (Kaluza, FlowJo, FCSExpress, SpectroFlo, FACSDiva)
Applications
We support multiparametric staining for cell subpopulation analysis in human and murine samples, including:
- Immune cells (Treg, Tnaive, Tconv, GC B cells, naïve B cells, TILs, TAMs)
- Cancer stem cells (ALDH, CD44/CD24 markers)
- Endothelial cells, myeloma cells, cardiac fibroblasts
- Cell subpopulations from spheroids, organoids, and co-cultures
Cell Cycle, Proliferation & Metabolic Analysis
- Cell cycle in mammalian and yeast cells (PI, Sytox, BrdU, EdU, FUCCI, Cyclins, Ki67)
- Glucose uptake studies (2NDBG)
- Lipids visualization (Bodipy, Nile Red for lipid droplets)
- Mitochondrial function (JC1, Mitotracker, Mitosox)
- Intracellular protein detection (TMR HaloTag)
- Reactive oxygen species detection (CellRox, DFCDA)
- ATP/ADP ratio monitoring (Perceval sensor)
- Cell death & apoptosis analysis (Caspase3, AnnV/PI, DRAQ7)
Genetic & Molecular Biology Applications
- Analysis of transduced/transfected/CRISPR-engineered cell lines
- Fluorescent protein detection (GFP, RFP, mCherry, Scarlet, Tomato, YFP…)
- Lineage tracking using RGB barcoding
- Protein interaction analysis (Duolink flowPLA® kit)
- Cell endocytosis studies (Dextran uptake)
- Microplastics uptake analysis in cell lines and murine tissues
- Quantification of antigen bound per cell (ABC) via QuantiBrite™ beads
Cell Sorting Capabilities
- Bulk sorting
- Sorting on specialized supports (e.g., glass slides for imaging, 96-well plates for 10X genomics, sequencing, mass spectrometry, high-content imaging)
- Single-cell sorting in 96- or 384-well plates
- Sorting with special precautions for metabolomics and NGS applications
Key users of the facility
Equipment
Analyzers: :
- FACSCantoII BD (3 lasers: V, B, R) - Details
- AttuneNxT ThermoFisher (4 lasers: V, B, Y, R) - Details
- Aurora Cytek (5 lasers: UV, V, B, Y, R) - Details
Sorters
Publications
PMID: 37713487
Clonal cooperation through soluble metabolite exchange facilitates metastatic outgrowth by modulating Allee effect.
Hershey BJ, Barozzi S, Orsenigo F, Pompei S, Iannelli F, Kamrad S, Matafora V, Pisati F, Calabrese L, Fragale G, Salvadori G, Martini E, Totaro MG, Magni S, Guan R, Parazzoli D, Maiuri P, Bachi A, Patil KR, Cosentino Lagomarsino M, Havas KM. Sci Adv. 2023 Sep 15;9(37):eadh4184. doi: 10.1126/sciadv.adh4184. Epub 2023 Sep 15. Free PMC article.
PMID: 37108076
Short-Term Exposure to Benzo(a)Pyrene Causes Disruption of GnRH Network in Zebrafish Embryos.
Gentile I, Vezzoli V, Martone S, Totaro MG, Bonomi M, Persani L, Marelli F. Int J Mol Sci. 2023 Apr 7;24(8):6913. doi: 10.3390/ijms24086913. Free PMC article.
PMID: 35918044
HDAC6 inhibition decreases leukemic stem cell expansion driven by Hedgehog hyperactivation by restoring primary ciliogenesis.
Pezzotta A, Gentile I, Genovese D, Totaro MG, Battaglia C, Leung AY, Fumagalli M, Parma M, Cazzaniga G, Fazio G, Alcalay M, Marozzi A, Pistocchi A. Pharmacol Res. 2022 Sep;183:106378. doi: 10.1016/j.phrs.2022.106378. Epub 2022 Jul 30.
PMID: 33653693
CD4 T Cell-Dependent Rejection of Beta-2 Microglobulin Null Mismatch Repair-Deficient Tumors.
Germano G, Lu S, Rospo G, Lamba S, Rousseau B, Fanelli S, Stenech D, Le DT, Hays J, Totaro MG, Amodio V, Chilà R, Mondino A, Diaz LA Jr, Di Nicolantonio F, Bardelli A. Cancer Discov. 2021 Jul;11(7):1844-1859. doi: 10.1158/2159-8290.CD-20-0987. Epub 2021 Mar 2.
PMID: 33436662
MITO-Luc/GFP zebrafish model to assess spatial and temporal evolution of cell proliferation in vivo.
de Latouliere L, Manni I, Ferrari L, Pisati F, Totaro MG, Gurtner A, Marra E, Pacello L, Pozzoli O, Aurisicchio L, Capogrossi MC, Deflorian G, Piaggio G. Sci Rep. 2021 Jan 12;11(1):671. PMID: 33436662 Free PMC article.
PMID: 33015043
HDAC8: A Promising Therapeutic Target for Acute Myeloid Leukemia.
Spreafico M, Gruszka AM, Valli D, Mazzola M, Deflorian G, Quintè A, Totaro MG, Battaglia C, Alcalay M, Marozzi A, Pistocchi A. Front Cell Dev Biol. 2020 Sep 4;8:844. doi: 10.3389/fcell.2020.00844. eCollection 2020. PMID: 33015043 Free PMC article.
PMID: 33903766
Heme catabolism by tumor-associated macrophages controls metastasis formation.
Consonni FM, Bleve A, Totaro MG, Storto M, Kunderfranco P, Termanini A, Pasqualini F, Alì C, Pandolfo C, Sgambelluri F, Grazia G, Santinami M, Maurichi A, Milione M, Erreni M, Doni A, Fabbri M, Gribaldo L, Rulli E, Soares MP, Torri V, Mortarini R, Anichini A, Sica A. Nat Immunol. 2021 May;22(5):595-606. doi: 10.1038/s41590-021-00921-5. Epub 2021 Apr 26.
PMID: 32265223
Tumor-Derived Prostaglandin E2 Promotes p50 NF-κB-Dependent Differentiation of Monocytic MDSCs.
Porta C, Consonni FM, Morlacchi S, Sangaletti S, Bleve A, Totaro MG, Larghi P, Rimoldi M, Tripodo C, Strauss L, Banfi S, Storto M, Pressiani T, Rimassa L, Tartari S, Ippolito A, Doni A, Soldà G, Duga S, Piccolo V, Ostuni R, Natoli G, Bronte V, Balzac F, Turco E, Hirsch E, Colombo MP, Sica A. Cancer Res. 2020 Jul 1;80(13):2874-2888. doi: 10.1158/0008-5472.CAN-19-2843. Epub 2020 Apr 7.
PMID: 18467122
Macrophage polarization in tumour progression.
Sica A, Larghi P, Mancino A, Rubino L, Porta C, Totaro MG, Rimoldi M, Biswas SK, Allavena P, Mantovani A. Semin Cancer Biol. 2008 Oct;18(5):349-55. doi: 10.1016/j.semcancer.2008.03.004. Epub 2008 Mar 26. Review.
PMID: 21995571
Macrophages in cancer and infectious diseases: the 'good' and the 'bad'.
Porta C, Riboldi E, Totaro MG, Strauss L, Sica A, Mantovani A. Immunotherapy. 2011 Oct;3(10):1185-202. doi: 10.2217/imt.11.116. Review.
PMID: 26267538
RORC1 Regulates Tumor-Promoting "Emergency" Granulo-Monocytopoiesis.
Strauss L, Sangaletti S, Consonni FM, Szebeni G, Morlacchi S, Totaro MG, Porta C, Anselmo A, Tartari S, Doni A, Zitelli F, Tripodo C, Colombo MP, Sica A. Cancer Cell. 2015 Aug 10;28(2):253-69. doi: 10.1016/j.ccell.2015.07.006. Free article.
PMID: 19616341
Cellular and molecular pathways linking inflammation and cancer.
Porta C, Larghi P, Rimoldi M, Totaro MG, Allavena P, Mantovani A, Sica A. Immunobiology. 2009;214(9-10):761-77. doi: 10.1016/j.imbio.2009.06.014. Epub 2009 Jul 17. Review.
PMID: 21948460
Origin and Functions of Tumor-Associated Myeloid Cells (TAMCs).
Sica A, Porta C, Morlacchi S, Banfi S, Strauss L, Rimoldi M, Totaro MG, Riboldi E. Cancer Microenviron. 2012 Aug;5(2):133-49. doi: 10.1007/s12307-011-0091-6. Epub 2011 Sep 24. Free PMC article.
PMID: 23049782
The p50 subunit of NF-κB orchestrates dendritic cell lifespan and activation of adaptive immunity.
Larghi P, Porta C, Riboldi E, Totaro MG, Carraro L, Orabona C, Sica A. PLoS One. 2012;7(9):e45279. doi: 10.1371/journal.pone.0045279. Epub 2012 Sep 25. Free PMC article.
PMID: 17825402
The histone H3 lysine-27 demethylase Jmjd3 links inflammation to inhibition of polycomb-mediated gene silencing.
De Santa F, Totaro MG, Prosperini E, Notarbartolo S, Testa G, Natoli G. Cell. 2007 Sep 21;130(6):1083-94. doi: 10.1016/j.cell.2007.08.019. Epub 2007 Sep 6. Free article.
PMID: 22127978
Enhanced interferon regulatory factor 3 binding to the interleukin-23p19 promoter correlates with enhanced interleukin-23 expression in systemic lupus erythematosus.
Smith S, Gabhann JN, Higgs R, Stacey K, Wahren-Herlenius M, Espinosa A, Totaro MG, Sica A, Ball E, Bell A, Johnston J, Browne P, O'Neill L, Kearns G, Jefferies CA. Arthritis Rheum. 2012 May;64(5):1601-9. doi: 10.1002/art.33494. Free article.
PMID: 18716661
The histone H3 lysine 27-specific demethylase Jmjd3 is required for neural commitment.
Burgold T, Spreafico F, De Santa F, Totaro MG, Prosperini E, Natoli G, Testa G. PLoS One. 2008 Aug 21;3(8):e3034. doi: 10.1371/journal.pone.0003034. Free PMC article.