Name

Citofluorimetry

Coordinator

Prof.ssa Daniela Fenoglio

Description

Flow cytometry is an extremely flexible technology platform for the characterization of cells or particles in suspension and allows to simultaneously analyze physical parameters and fluorescence characteristics of many hundreds of cells or particles every second. According to a more modern meaning, flow cytometry is the technology that allows to evaluate qualitatively and quantitatively the molecules expressed on the surface or inside the cell, allowing a precise identification of the cell type in question. Moreover, using multiplex assays can simultaneously measure multiple analytes (cytokines, chemokines, inflammatory mediators, etc.) in a single sample with cytometry applications. The technology platform can be applied in many fields of biology and medicine.

Cytometry Facility operates in two areas: experimental and clinical cytometry.

Purpose of experimental cytometry are : a) to provide support to the design and execution of research projects in the biomedical field, with particular reference to the development of new analytical protocols; b) to provide a service to internal and external research groups, including collaboration with small and medium-size companies, c) to support projects aimed at the development of innovative protocols of therapeutic strategies in preclinical models or in vitro evaluation and in vivo effects of drugs, cytokines or compounds of various nature of primary cells or continuous cell lines; d) to support the development of industrial products involving the use of flow cytometry.

Clinical cytometry has aimed to study the phenotypic and functional cells of the immune system in health and disease (autoimmunity, cancer, infectious diseases, immunodeficiency). In addition, the service offers optimized protocols of cytometric cell analyses for immunological monitoring in preclinical and clinical trials according to international guidelines established by the Cancer Immunotherapy Consortium of the Cancer Research Institute and the 'Association for Cancer Immunotherapy CIMT Immunoguiding Program.

The Flow Cytometry Facility is equipped with the following cell analyzers:
a) BD Facs Canto II cell analyzer, : 3 laser (488 nm, 633 nm, 405 nm), 10 parameters (FSC, SSC and 8 fluorescence colors);
b) BD LSRFortessa X20 cell analyzer: 4 laser (488 nm, 640 nm, 405 nm, 561 nm), 18 parameters (FSC, SSC and 8 fluorescence colors).

Data analysis is performed using the following softwares: DIVA V6.1.3, DIVA v8.1.0, ModFit LT v3.2, FlowJo v7.6.1, FCAPArray v1.0.1.

Moreover, Flow Cytometry Facility is active in continuing education in the field cytometric by Training of users in the use of analytical cytometers and assisting researchers with analysis and interpretation of cytometric data.

Applications:

• Immunophenotyping of murine and human cells
• Apoptosis and necrosis
• Proliferation
• Cytoxicity
• Evaluation of cytokine profile by intracellular staining
• Analysis of antigen-specific CD8+T cells by labelled multimer staining
• Evaluation of degranulation of basophylic granulocytes
• Quantitative detection of multiple analytes by bead-based reagents
• Cell viability assays
• Oxidative stress
• Quantification and analysis of microvescicles