{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,26]],"date-time":"2026-10-26T23:18:41Z","timestamp":1793056721691,"version":"4.5.0"},"reference-count":51,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2021,1,29]],"date-time":"2021-01-29T00:00:00Z","timestamp":1611878400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":["frontiersin.org"],"crossmark-restriction":true},"short-container-title":["Front. Mol. Biosci."],"abstract":"<jats:p>The Nobel Prize-deserving concept of blocking inhibitory pathways in T cells, to unleash their anti-tumoral capacity, became one of the pillars of cancer treatment in the last decade and has resulted in durable clinical responses for multiple cancer types. Currently, two of the most important goals in cancer immunotherapy are to understand the mechanisms resulting in failure to checkpoint blockade and to identify predictive immunological biomarkers that correlate to treatment response, disease progression or adverse effects. The identification and validation of biomarkers for routine clinical use is not only critical to monitor disease or treatment progression, but also to personalize and develop new therapies. To achieve these goals, powerful research tools are needed. Flow cytometry stands as one of the most successful single-cell analytical tools used to characterize immune cell phenotypes to monitor solid tumors, hematological malignancies, minimal residual disease or metastatic progression. This technology has been fundamental in diagnosis, treatment and translational research in cancer clinical trials. Most recently, the need to evaluate simultaneously more features in each cell has pushed the field to implement more powerful adaptations beyond conventional flow cytometry, including Full Spectrum Flow Cytometry (FSFC). FSFC captures the full emission spectrum of fluorescent molecules using arrays of highly sensitive light detectors, and to date has enabled characterization of 40 parameters in a single sample. We will summarize the contributions of this technology to the advancement of research in immunotherapy studies and discuss best practices to obtain reliable, robust and reproducible FSFC results.<\/jats:p>","DOI":"10.3389\/fmolb.2020.612801","type":"journal-article","created":{"date-parts":[[2021,1,29]],"date-time":"2021-01-29T04:34:31Z","timestamp":1611894871000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":99,"title":["Full Spectrum Flow Cytometry as a Powerful Technology for Cancer Immunotherapy Research"],"prefix":"10.3389","volume":"7","author":[{"given":"Diana L.","family":"Bonilla","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Gil","family":"Reinin","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Edmond","family":"Chua","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1965","published-online":{"date-parts":[[2021,1,29]]},"reference":[{"key":"B1","doi-asserted-by":"publisher","first-page":"1202","DOI":"10.1016\/j.cell.2015.08.047","article-title":"Checkpoints","volume":"162","author":"Allison","year":"2015","journal-title":"Cell"},{"key":"B2","doi-asserted-by":"publisher","first-page":"127","DOI":"10.1038\/s41586-020-2403-9","article-title":"Senolytic CAR T cells reverse senescence-associated pathologies","volume":"583","author":"Amor","year":"2020","journal-title":"Nature"},{"key":"B3","doi-asserted-by":"publisher","first-page":"1","DOI":"10.1002\/cpim.37","article-title":"High-dimensional fluorescence cytometry","volume":"119","author":"Ashhurst","year":"2017","journal-title":"Curr. Protocols Immunol"},{"key":"B4","doi-asserted-by":"publisher","DOI":"10.1101\/2020.07.31.190454","article-title":"Unsupervised machine learning reveals key immune cell subsets in COVID-19, rhinovirus infection, and cancer therapy","author":"Barone","year":"2020","journal-title":"bioRxiv"},{"key":"B5","doi-asserted-by":"publisher","first-page":"13","DOI":"10.1007\/978-1-4939-3302-0_2","article-title":"Principles of amnis imaging flow cytometry","volume":"1389","author":"Basiji","year":"2016","journal-title":"Methods Mol. Biol"},{"key":"B6","doi-asserted-by":"publisher","first-page":"1537","DOI":"10.3390\/cancers12061537","article-title":"Phase I study of ficlatuzumab and cetuximab in cetuximab-resistant, recurrent\/metastatic head and neck cancer","volume":"12","author":"Bauman","year":"2020","journal-title":"Cancers"},{"key":"B7","doi-asserted-by":"publisher","first-page":"639","DOI":"10.1038\/nbt.2283","article-title":"From single cells to deep phenotypes in cancer","volume":"30","author":"Bendall","year":"2012","journal-title":"Nat. Biotechnol"},{"key":"B8","doi-asserted-by":"publisher","first-page":"182","DOI":"10.3390\/vaccines8020182","article-title":"Tumor membrane vesicle vaccine augments the efficacy of anti-PD1 antibody in immune checkpoint inhibitor-resistant squamous cell carcinoma models of head and neck cancer","volume":"8","author":"Bommireddy","year":"2020","journal-title":"Vaccine"},{"key":"B9","doi-asserted-by":"publisher","first-page":"184","DOI":"10.1038\/s43018-020-0028-4","article-title":"Extracellular cGAMP is a cancer-cell-produced immunotransmitter involved in radiation-induced anticancer immunity","volume":"1","author":"Carozza","year":"2020","journal-title":"Nat. Cancer"},{"key":"B10","doi-asserted-by":"publisher","first-page":"1457","DOI":"10.1002\/eji.201970107","article-title":"Guidelines for the use of flow cytometry and cell sorting in immunological studies (second edition)","volume":"49","author":"Cossarizza","year":"2019","journal-title":"Eur. J. Immunol"},{"key":"B11","doi-asserted-by":"publisher","first-page":"114","DOI":"10.14440\/jbm.2019.283","article-title":"Detection of clinically relevant immune checkpoint markers by multicolor flow cytometry","volume":"6","author":"Cunningham","year":"2019","journal-title":"J. Biol. Methods"},{"key":"B12","doi-asserted-by":"publisher","first-page":"824","DOI":"10.1002\/cyto.a.24016","article-title":"High-dimensional data analysis algorithms yield comparable results for mass cytometry and spectral flow cytometry data","volume":"97","author":"Ferrer-Font","year":"","journal-title":"Cytometry A"},{"key":"B13","doi-asserted-by":"publisher","first-page":"e70","DOI":"10.1002\/cpcy.70","article-title":"Panel design and optimization for high-dimensional immunophenotyping assays using spectral flow cytometry","volume":"92","author":"Ferrer-Font","year":"","journal-title":"Curr. Protoc. Cytom"},{"key":"B14","doi-asserted-by":"publisher","first-page":"e74","DOI":"10.1002\/cpcy.74","article-title":"Acquisition of high\u2010quality spectral flow cytometry data","volume":"93","author":"Fox","year":"2020","journal-title":"Curr. Protoc. Cytom"},{"key":"B15","doi-asserted-by":"publisher","first-page":"214","DOI":"10.1186\/1479-5876-9-214","article-title":"Defining the critical hurdles in cancer immunotherapy","volume":"9","author":"Fox","year":"2011","journal-title":"J. Transl. Med"},{"key":"B16","doi-asserted-by":"publisher","first-page":"1","DOI":"10.1016\/bs.acr.2019.03.001","article-title":"Translation of cancer immunotherapy from the bench to the bedside","volume":"143","author":"Guo","year":"2019","journal-title":"Adv. Cancer Res"},{"key":"B17","doi-asserted-by":"publisher","first-page":"95","DOI":"10.1186\/s13045-020-00928-9","article-title":"DT2216-a Bcl-xL-specific degrader is highly active against Bcl-xL-dependent T cell lymphomas","volume":"13","author":"He","year":"2020","journal-title":"J. Hematol. Oncol"},{"key":"B18","doi-asserted-by":"publisher","first-page":"193","DOI":"10.1016\/j.blre.2013.06.005","article-title":"Developments in the immunophenotypic analysis of haematological malignancies","volume":"27","author":"Heel","year":"2013","journal-title":"Blood Rev"},{"key":"B19","doi-asserted-by":"publisher","first-page":"711","DOI":"10.1056\/NEJMoa1003466","article-title":"Improved survival with ipilimumab in patients with metastatic melanoma","volume":"363","author":"Hodi","year":"2010","journal-title":"N. Engl. J. Med"},{"key":"B20","doi-asserted-by":"publisher","first-page":"1","DOI":"10.1007\/82_2014_367","article-title":"High-dimensional single-cell cancer biology","volume":"377","author":"Irish","year":"2014","journal-title":"Curr. Top. Microbiol. Immunol"},{"key":"B21","doi-asserted-by":"publisher","first-page":"2295","DOI":"10.4049\/jimmunol.1900778","article-title":"Cancer cell-intrinsic expression of MHC class II regulates the immune microenvironment and response to Anti-PD-1 therapy in lung adenocarcinoma","volume":"204","author":"Johnson","year":"2020","journal-title":"J. Immunol"},{"key":"B22","doi-asserted-by":"publisher","first-page":"116","DOI":"10.1002\/cyto.a.23882","article-title":"Rigor and reproducibility of cytometry practices for immuno-oncology: a multifaceted challenge","volume":"97","author":"Laskowski","year":"2020","journal-title":"Cytom. A"},{"key":"B23","doi-asserted-by":"publisher","first-page":"5397","DOI":"10.1172\/JCI133008","article-title":"Graft IL-33 regulates infiltrating macrophages to protect against chronic rejection","volume":"130","author":"Li","year":"2020","journal-title":"J. Clin. Investig"},{"key":"B24","doi-asserted-by":"publisher","first-page":"2457","DOI":"10.1681\/ASN.2020030320","article-title":"Coordinated circulating T follicular helper and activated B cell responses underlie the onset of antibody-mediated rejection in kidney transplantation","volume":"31","author":"Louis","year":"2020","journal-title":"J. Am. Soc. Nephrol"},{"key":"B25","doi-asserted-by":"publisher","first-page":"44","DOI":"10.1186\/s40425-015-0085-x","article-title":"Immune monitoring technology primer: flow and mass cytometry","volume":"3","author":"Maecker","year":"2015","journal-title":"J. Immunother. Cancer"},{"key":"B26","doi-asserted-by":"publisher","first-page":"217","DOI":"10.2991\/jegh.k.191008.001","article-title":"Current cancer epidemiology","volume":"9","author":"Mattiuzzi","year":"2019","journal-title":"JEGH"},{"key":"B27","volume-title":"Translational analysis from SCALOP trial: CCL5 as a prognostic biomarker and a potentially actionable target in locally advanced pancreatic cancer (LAPC)","author":"Mukherjee","year":"2020"},{"key":"B28","doi-asserted-by":"publisher","first-page":"e000987","DOI":"10.1136\/jitc-2020-000987","article-title":"Immunohistochemical scoring of CD38 in the tumor microenvironment predicts responsiveness to anti-PD-1\/PD-L1 immunotherapy in hepatocellular carcinoma","volume":"8","author":"Ng","year":"2020","journal-title":"J. Immunother. Cancer"},{"key":"B29","doi-asserted-by":"publisher","first-page":"1165","DOI":"10.1002\/cyto.a.24211","article-title":"Evaluating spectral cytometry for immune profiling in viral disease","volume":"97","author":"Niewold","year":"2020","journal-title":"Cytom. A"},{"key":"B30","doi-asserted-by":"publisher","first-page":"28","DOI":"10.1002\/0471142956.cy0127s63","article-title":"Spectral flow cytometry","volume":"63","author":"Nolan","year":"2013","journal-title":"Curr. Protoc. Cytom"},{"key":"B31","doi-asserted-by":"publisher","first-page":"253","DOI":"10.1002\/cyto.a.22241","article-title":"Visible and near infrared fluorescence spectral flow cytometry","author":"Nolan","year":"2013","journal-title":"Cytom. A"},{"key":"B32","doi-asserted-by":"publisher","first-page":"3533","DOI":"10.1038\/s41467-018-05886-y","article-title":"Loss-of-function mutations in ATP6AP1 and ATP6AP2 in granular cell tumors","volume":"9","author":"Pareja","year":"2018","journal-title":"Nat. Commun"},{"key":"B33","doi-asserted-by":"publisher","first-page":"1044","DOI":"10.1002\/cyto.a.24213","article-title":"OMIP-069: Forty-color full spectrum flow cytometry panel for deep immunophenotyping of major cell subsets in human peripheral blood","volume":"97","author":"Park","year":"2020","journal-title":"Cytom. A"},{"key":"B35","doi-asserted-by":"publisher","first-page":"36","DOI":"10.1002\/cyto.a.20889","article-title":"Multispectral cytometry: the next generation","volume":"11","author":"Robinson","year":"2004","journal-title":"Biophotonics Int"},{"key":"B36","doi-asserted-by":"publisher","first-page":"823","DOI":"10.1002\/cyto.a.23779","article-title":"Spectral flow cytometry- Quo vadimus ?","volume":"95","author":"Robinson","year":"2019","journal-title":"Cytom. A"},{"key":"B53","doi-asserted-by":"publisher","first-page":"169","DOI":"10.1016\/j.cell.2020.11.029","article-title":"Functional SARS-CoV-2-specific immune memory persists after mild COVID-19","volume":"184","author":"Rodda","year":"2021","journal-title":"Cell"},{"key":"B37","doi-asserted-by":"publisher","first-page":"205","DOI":"10.1016\/j.cell.2015.03.030","article-title":"Immune checkpoint targeting in cancer therapy: toward combination strategies with curative potential","volume":"161","author":"Sharma","year":"2015","journal-title":"Cell"},{"key":"B38","doi-asserted-by":"publisher","first-page":"805","DOI":"10.1038\/nrc3153","article-title":"Novel cancer immunotherapy agents with survival benefit: recent successes and next steps","volume":"11","author":"Sharma","year":"2011","journal-title":"Nat. Rev. Cancer"},{"key":"B39","doi-asserted-by":"publisher","first-page":"1","DOI":"10.1038\/s41598-020-65129-3","article-title":"thymic resident nKt cell subsets show differential requirements for CD28 co-stimulation during antigenic activation","volume":"10","author":"Shissler","year":"2020","journal-title":"Sci. Rep"},{"key":"B40","doi-asserted-by":"publisher","first-page":"eaba0995","DOI":"10.1126\/sciadv.aba0995","article-title":"Adjuvant-free nanofiber vaccine induces in situ lung dendritic cell activation and TH17 responses","volume":"6","author":"Si","year":"2020","journal-title":"Sci. Adv"},{"key":"B41","doi-asserted-by":"publisher","first-page":"1401","DOI":"10.1016\/j.cell.2020.08.002","article-title":"Elevated calprotectin and abnormal myeloid cell subsets discriminate severe from Mild COVID-19","volume":"182","author":"Silvin","year":"2020","journal-title":"Cell"},{"key":"B42","doi-asserted-by":"publisher","first-page":"1057","DOI":"10.1002\/cyto.a.24041","article-title":"Phenotypic analysis of the mouse hematopoietic hierarchy using spectral cytometry: from stem cell subsets to early progenitor compartments","volume":"97","author":"Solomon","year":"2020","journal-title":"Cytom. A"},{"key":"B43","doi-asserted-by":"publisher","first-page":"780","DOI":"10.1016\/j.cell.2016.04.019","article-title":"Mass cytometry: single cells, many features","volume":"165","author":"Spitzer","year":"2016","journal-title":"Cell"},{"key":"B44","doi-asserted-by":"publisher","first-page":"865","DOI":"10.1038\/nmeth.4380","article-title":"Simultaneous epitope and transcriptome measurement in single cells","volume":"14","author":"Stoeckius","year":"2017","journal-title":"Nat. Methods"},{"key":"B45","doi-asserted-by":"publisher","first-page":"1069","DOI":"10.1158\/2159-8290.CD-18-0367","article-title":"Fundamental mechanisms of immune checkpoint blockade therapy","volume":"8","author":"Wei","year":"2018","journal-title":"Cancer Discov"},{"key":"B46","doi-asserted-by":"publisher","first-page":"111","DOI":"10.4137\/CIN.S16346","article-title":"Managing multi-center flow cytometry data for immune monitoring","volume":"13","author":"White","year":"2014","journal-title":"Cancer Inf"},{"key":"B47","doi-asserted-by":"publisher","first-page":"4244","DOI":"10.1182\/bloodadvances.2020002355","article-title":"Charge-altering releasable transporters enable phenotypic manipulation of natural killer cells for cancer immunotherapy","volume":"4","author":"Wilk","year":"2020","journal-title":"Blood Adv"},{"key":"B48","doi-asserted-by":"publisher","first-page":"1708064","DOI":"10.1080\/2162402X.2019.1708064","article-title":"Checkpoint molecules coordinately restrain hyperactivated effector T cells in the tumor microenvironment","volume":"9","author":"Yang","year":"2020","journal-title":"Oncoimmunology"},{"key":"B49","doi-asserted-by":"publisher","first-page":"3","DOI":"10.1186\/s40425-016-0107-3","article-title":"Novel technologies and emerging biomarkers for personalized cancer immunotherapy","volume":"4","author":"Yuan","year":"2016","journal-title":"J. Immunother. Cancer"},{"key":"B50","doi-asserted-by":"publisher","first-page":"112186","DOI":"10.1016\/j.ejmech.2020.112186","article-title":"Discovery of PROTAC BCL-XL degraders as potent anticancer agents with low on-target platelet toxicity","volume":"192","author":"Zhang","year":"2020","journal-title":"Eur. J. Med. Chem"},{"key":"B51","doi-asserted-by":"publisher","first-page":"6054","DOI":"10.1172\/JCI138538","article-title":"Boosting NAD level suppresses inflammatory activation of PBMC in heart failure","volume":"130","author":"Zhou","year":"2020","journal-title":"J. Clin. Invest"}],"container-title":["Frontiers in Molecular Biosciences"],"original-title":[],"link":[{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fmolb.2020.612801\/full","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2021,1,29]],"date-time":"2021-01-29T04:34:33Z","timestamp":1611894873000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fmolb.2020.612801\/full"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2021,1,29]]},"references-count":51,"alternative-id":["10.3389\/fmolb.2020.612801"],"URL":"https:\/\/doi.org\/10.3389\/fmolb.2020.612801","relation":{},"ISSN":["2296-889X"],"issn-type":[{"value":"2296-889X","type":"electronic"}],"subject":[],"published":{"date-parts":[[2021,1,29]]},"article-number":"612801"}}