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. 2020 Jul 14;29:0963689720923578. doi: 10.1177/0963689720923578

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© The Author(s) 2020

This article is distributed under the terms of the Creative Commons Attribution-NonCommercial 4.0 License (https://creativecommons.org/licenses/by-nc/4.0/) which permits non-commercial use, reproduction and distribution of the work without further permission provided the original work is attributed as specified on the SAGE and Open Access pages (https://us.sagepub.com/en-us/nam/open-access-at-sage).

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Figure 4. — Example of flow cytometry gating strategy for phenotyping of Tregs in both flask and Quantum system Treg-harvested cells. Treg frequency is based on the number of CD4⁺CD25⁺FoxP3⁺ cells within the CD25⁺FoxP3⁺ double-positive gate that are above background FMO fluorescence for both FoxP3 and CD25. Flask gate labels, plots left to right: SSC vs FSC cells, singlets (SSC-H vs SSC-A), CD3⁺ dead and CD3⁺ live, CD8⁺ vs CD4⁺, CD25⁺FoxP3⁺ Tregs; Quantum system gate labels, plots left to right: SSC vs FSC cells, singlets (SSC-H vs SSC-A), CD3⁺ dead and CD3⁺ live, CD8⁺ vs CD4⁺, CD25⁺FoxP3⁺ Tregs. FoxP3 AF: 647; CD8 BUV: 737; CD3 BV: 786; CD4: BUV395; FMO: fluorescence minus one; FoxP3: forkhead box protein P3; FSC: forward scatter; PE: CD25; SSC: side scatter; SSC-A: side scatter - area; SSC-H: side scatter - height; Tregs: regulatory T cells.