Journal
CANCER IMMUNOLOGY IMMUNOTHERAPY
Volume 64, Issue 10, Pages 1271-1286Publisher
SPRINGER
DOI: 10.1007/s00262-015-1729-x
Keywords
Consensus; Tregs; Monitoring; Flow cytometry; Phenotyping
Categories
Funding
- Wallace H. Coulter Foundation (Miami, Florida, USA)
- Bontius stichting
- Wallace Coulter Foundation
- Deutsche Forschungsgemeinschaft [SFB 685/Z5, Sonderforschungsbereich 633, Sonderforschungsbereich 650]
- Norwegian Cancer Society
- Research Council of Norway
- Kristian Gerhard Jebsen Foundation
- Wellcome Trust [086983/Z/08/Z]
- Cancer Research Wales
- Nijmegen Institute for Infection, Inflammation, and Immunity [151692]
- Division of Intramural Research, National Institute of Allergy and Infectious Diseases, National Institute of Health (DIR/NIAID/NIH)
- Duke University Center for AIDS Research (CFAR), an National Institute of Health [5P30 AI064518]
- Dutch Cancer Society [2009-4400]
- National Institute of Health [R01-CA168628]
- Wellcome Trust [086983/Z/08/Z] Funding Source: Wellcome Trust
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Regulatory T cell (Treg)-mediated immunosuppression is considered a major obstacle for successful cancer immunotherapy. The association between clinical outcome and Tregs is being studied extensively in clinical trials, but unfortunately, no consensus has been reached about (a) the markers and (b) the gating strategy required to define human Tregs in this context, making it difficult to draw final conclusions. Therefore, we have organized an international workshop on the detection and functional testing of Tregs with leading experts in the field, and 40 participants discussing different analyses and the importance of different markers and context in which Tregs were analyzed. This resulted in a rationally composed ranking list of Treg markers. Subsequently, the proposed Treg markers were tested to get insight into the overlap/differences between the most frequently used Treg definitions and their utility for Treg detection in various human tissues. Here, we conclude that the CD3, CD4, CD25, CD127, and FoxP3 markers are the minimally required markers to define human Treg cells. Staining for Ki67 and CD45RA showed to provide additional information on the activation status of Tregs. The use of markers was validated in a series of PBMC from healthy donors and cancer patients, as well as in tumor-draining lymph nodes and freshly isolated tumors. In conclusion, we propose an essential marker set comprising antibodies to CD3, CD4, CD25, CD127, Foxp3, Ki67, and CD45RA and a corresponding robust gating strategy for the context-dependent analysis of Tregs by flow cytometry.
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