Multicentre Harmonisation of a Six-Colour Flow Cytometry Panel for Naïve/Memory T Cell Immunomonitoring
Background. Personalised medicine in oncology needs standardised immunological assays. Flow cytometry (FCM) methods represent an essential tool for immunomonitoring, and their harmonisation is crucial to obtain comparable data in multicentre clinical trials. The objective of this study was to design...
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2020-01-01
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| Series: | Journal of Immunology Research |
| Online Access: | http://dx.doi.org/10.1155/2020/1938704 |
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| author | Iole Macchia Valentina La Sorsa Irene Ruspantini Massimo Sanchez Valentina Tirelli Maria Carollo Giorgio Fedele Pasqualina Leone Giovanna Schiavoni Carla Buccione Paola Rizza Paola Nisticò Belinda Palermo Stefania Morrone Helena Stabile Aurelia Rughetti Marianna Nuti Ilaria Grazia Zizzari Cinzia Fionda Roberta Maggio Cristina Capuano Concetta Quintarelli Matilde Sinibaldi Chiara Agrati Rita Casetti Andrea Rozo Gonzalez Floriana Iacobone Angela Gismondi Filippo Belardelli Mauro Biffoni Francesca Urbani |
| author_facet | Iole Macchia Valentina La Sorsa Irene Ruspantini Massimo Sanchez Valentina Tirelli Maria Carollo Giorgio Fedele Pasqualina Leone Giovanna Schiavoni Carla Buccione Paola Rizza Paola Nisticò Belinda Palermo Stefania Morrone Helena Stabile Aurelia Rughetti Marianna Nuti Ilaria Grazia Zizzari Cinzia Fionda Roberta Maggio Cristina Capuano Concetta Quintarelli Matilde Sinibaldi Chiara Agrati Rita Casetti Andrea Rozo Gonzalez Floriana Iacobone Angela Gismondi Filippo Belardelli Mauro Biffoni Francesca Urbani |
| author_sort | Iole Macchia |
| collection | DOAJ |
| description | Background. Personalised medicine in oncology needs standardised immunological assays. Flow cytometry (FCM) methods represent an essential tool for immunomonitoring, and their harmonisation is crucial to obtain comparable data in multicentre clinical trials. The objective of this study was to design a harmonisation workflow able to address the most effective issues contributing to intra- and interoperator variabilities in a multicentre project. Methods. The Italian National Institute of Health (Istituto Superiore di Sanità, ISS) managed a multiparametric flow cytometric panel harmonisation among thirteen operators belonging to five clinical and research centres of Lazio region (Italy). The panel was based on a backbone mixture of dried antibodies (anti-CD3, anti-CD4, anti-CD8, anti-CD45RA, and anti-CCR7) to detect naïve/memory T cells, recognised as potential prognostic/predictive immunological biomarkers in cancer immunotherapies. The coordinating centre distributed frozen peripheral blood mononuclear cells (PBMCs) and fresh whole blood (WB) samples from healthy donors, reagents, and Standard Operating Procedures (SOPs) to participants who performed experiments by their own equipment, in order to mimic a real-life scenario. Operators returned raw and locally analysed data to ISS for central analysis and statistical elaboration. Results. Harmonised and reproducible results were obtained by sharing experimental set-up and procedures along with centralising data analysis, leading to a reduction of cross-centre variability for naïve/memory subset frequencies particularly in the whole blood setting. Conclusion. Our experimental and analytical working process proved to be suitable for the harmonisation of FCM assays in a multicentre setting, where high-quality data are required to evaluate potential immunological markers, which may contribute to select better therapeutic options. |
| format | Article |
| id | doaj-art-04486219890645d4a2e10952819dce83 |
| institution | Kabale University |
| issn | 2314-8861 2314-7156 |
| language | English |
| publishDate | 2020-01-01 |
| publisher | Wiley |
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| series | Journal of Immunology Research |
| spelling | doaj-art-04486219890645d4a2e10952819dce832025-02-03T05:49:31ZengWileyJournal of Immunology Research2314-88612314-71562020-01-01202010.1155/2020/19387041938704Multicentre Harmonisation of a Six-Colour Flow Cytometry Panel for Naïve/Memory T Cell ImmunomonitoringIole Macchia0Valentina La Sorsa1Irene Ruspantini2Massimo Sanchez3Valentina Tirelli4Maria Carollo5Giorgio Fedele6Pasqualina Leone7Giovanna Schiavoni8Carla Buccione9Paola Rizza10Paola Nisticò11Belinda Palermo12Stefania Morrone13Helena Stabile14Aurelia Rughetti15Marianna Nuti16Ilaria Grazia Zizzari17Cinzia Fionda18Roberta Maggio19Cristina Capuano20Concetta Quintarelli21Matilde Sinibaldi22Chiara Agrati23Rita Casetti24Andrea Rozo Gonzalez25Floriana Iacobone26Angela Gismondi27Filippo Belardelli28Mauro Biffoni29Francesca Urbani30Department of Oncology and Molecular Medicine, Istituto Superiore di Sanità (ISS), Rome 00161, ItalyResearch Coordination and Support Service, CoRI, ISS, Rome 00161, ItalyCore Facilities, ISS, Rome 00161, ItalyCore Facilities, ISS, Rome 00161, ItalyCore Facilities, ISS, Rome 00161, ItalyCore Facilities, ISS, Rome 00161, ItalyDepartment of Infectious Diseases, ISS, Rome 00161, ItalyDepartment of Infectious Diseases, ISS, Rome 00161, ItalyDepartment of Oncology and Molecular Medicine, Istituto Superiore di Sanità (ISS), Rome 00161, ItalyDepartment of Oncology and Molecular Medicine, Istituto Superiore di Sanità (ISS), Rome 00161, ItalyCenter for Gender-Specific Medicine, ISS, Rome 00161, ItalyUnit of Tumor Immunology and Immunotherapy, IRCCS Regina Elena National Cancer Institute (IRE), Rome 00128, ItalyUnit of Tumor Immunology and Immunotherapy, IRCCS Regina Elena National Cancer Institute (IRE), Rome 00128, ItalyDepartment of Experimental Medicine, Sapienza University of Rome (SUR), 00185, ItalyDepartment of Molecular Medicine, SUR, Rome 00185, ItalyDepartment of Experimental Medicine, Sapienza University of Rome (SUR), 00185, ItalyDepartment of Experimental Medicine, Sapienza University of Rome (SUR), 00185, ItalyDepartment of Experimental Medicine, Sapienza University of Rome (SUR), 00185, ItalyDepartment of Molecular Medicine, SUR, Rome 00185, ItalyClinical Research, Imperial College, London W12 ONN, UKDepartment of Experimental Medicine, Sapienza University of Rome (SUR), 00185, ItalyOnco-Hematology Department, IRCCS Bambino Gesù Children’s Hospital (OPBG), Rome 00165, ItalyOnco-Hematology Department, IRCCS Bambino Gesù Children’s Hospital (OPBG), Rome 00165, ItalyCellular Immunology Laboratory, IRCCS National Institute for Infectious Diseases “L. Spallanzani” (INMI), Rome 00149, ItalyCellular Immunology Laboratory, IRCCS National Institute for Infectious Diseases “L. Spallanzani” (INMI), Rome 00149, ItalyDepartment of Oncology and Molecular Medicine, Istituto Superiore di Sanità (ISS), Rome 00161, ItalyDepartment of Oncology and Molecular Medicine, Istituto Superiore di Sanità (ISS), Rome 00161, ItalyDepartment of Molecular Medicine, SUR, Rome 00185, ItalyDepartment of Oncology and Molecular Medicine, Istituto Superiore di Sanità (ISS), Rome 00161, ItalyDepartment of Oncology and Molecular Medicine, Istituto Superiore di Sanità (ISS), Rome 00161, ItalyDepartment of Oncology and Molecular Medicine, Istituto Superiore di Sanità (ISS), Rome 00161, ItalyBackground. Personalised medicine in oncology needs standardised immunological assays. Flow cytometry (FCM) methods represent an essential tool for immunomonitoring, and their harmonisation is crucial to obtain comparable data in multicentre clinical trials. The objective of this study was to design a harmonisation workflow able to address the most effective issues contributing to intra- and interoperator variabilities in a multicentre project. Methods. The Italian National Institute of Health (Istituto Superiore di Sanità, ISS) managed a multiparametric flow cytometric panel harmonisation among thirteen operators belonging to five clinical and research centres of Lazio region (Italy). The panel was based on a backbone mixture of dried antibodies (anti-CD3, anti-CD4, anti-CD8, anti-CD45RA, and anti-CCR7) to detect naïve/memory T cells, recognised as potential prognostic/predictive immunological biomarkers in cancer immunotherapies. The coordinating centre distributed frozen peripheral blood mononuclear cells (PBMCs) and fresh whole blood (WB) samples from healthy donors, reagents, and Standard Operating Procedures (SOPs) to participants who performed experiments by their own equipment, in order to mimic a real-life scenario. Operators returned raw and locally analysed data to ISS for central analysis and statistical elaboration. Results. Harmonised and reproducible results were obtained by sharing experimental set-up and procedures along with centralising data analysis, leading to a reduction of cross-centre variability for naïve/memory subset frequencies particularly in the whole blood setting. Conclusion. Our experimental and analytical working process proved to be suitable for the harmonisation of FCM assays in a multicentre setting, where high-quality data are required to evaluate potential immunological markers, which may contribute to select better therapeutic options.http://dx.doi.org/10.1155/2020/1938704 |
| spellingShingle | Iole Macchia Valentina La Sorsa Irene Ruspantini Massimo Sanchez Valentina Tirelli Maria Carollo Giorgio Fedele Pasqualina Leone Giovanna Schiavoni Carla Buccione Paola Rizza Paola Nisticò Belinda Palermo Stefania Morrone Helena Stabile Aurelia Rughetti Marianna Nuti Ilaria Grazia Zizzari Cinzia Fionda Roberta Maggio Cristina Capuano Concetta Quintarelli Matilde Sinibaldi Chiara Agrati Rita Casetti Andrea Rozo Gonzalez Floriana Iacobone Angela Gismondi Filippo Belardelli Mauro Biffoni Francesca Urbani Multicentre Harmonisation of a Six-Colour Flow Cytometry Panel for Naïve/Memory T Cell Immunomonitoring Journal of Immunology Research |
| title | Multicentre Harmonisation of a Six-Colour Flow Cytometry Panel for Naïve/Memory T Cell Immunomonitoring |
| title_full | Multicentre Harmonisation of a Six-Colour Flow Cytometry Panel for Naïve/Memory T Cell Immunomonitoring |
| title_fullStr | Multicentre Harmonisation of a Six-Colour Flow Cytometry Panel for Naïve/Memory T Cell Immunomonitoring |
| title_full_unstemmed | Multicentre Harmonisation of a Six-Colour Flow Cytometry Panel for Naïve/Memory T Cell Immunomonitoring |
| title_short | Multicentre Harmonisation of a Six-Colour Flow Cytometry Panel for Naïve/Memory T Cell Immunomonitoring |
| title_sort | multicentre harmonisation of a six colour flow cytometry panel for naive memory t cell immunomonitoring |
| url | http://dx.doi.org/10.1155/2020/1938704 |
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