Detailed Treg Cell Identification

Flow cytometry was performed on the Gallios 10/3 flow cytometer and the results were analyzed by Kaluza Analysis Software (Both from Beckman Coulter). Preparation of cells was performed as described previously (Bruck et al., 2022 (link); Klasen et al., 2019 (link); Kofler et al., 2006 (link)). In brief, cells were stained with cell surface targets including CD4 (Cat. No. 347413), CD25 (Cat. No. 340907), and CD127 (Cat. No. 566399), then fixed and permeabilized by the BD Cytofix/Cytoperm Kit (BD Bioscience) or the True‐NuclearTM Transcription Factor Buffer Set (BioLegend Inc.) according to the manufacturer's instructions. Anti‐FoxP3 antibodies (Cat. No. 320116) and isotype antibodies (Cat. No. 981812) were purchased from Bio Legend Inc. Dead cells were excluded by the LIVE/DEADTM Fixable Dead Cell Stain Kit (Invitrogen, Thermo Fisher Scientific). Treg cells were defined as CD4⁺CD25^highCD127^lowFoxP3⁺ T cells.