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Subsequently, these cells were washed with cold PBS and Crenolanib purchase with permeabilization buffer (eBioscience, San Diego, CA, USA) and then 20?��L anti-human FoxP3-PE (BD Biosciences, San Jose, CA,USA) was added and the cells were incubated at 4��C for 30?min. Stained cells were analyzed with a FACSAria II flow cytometer (BD Biosciences, San Jose, CA, USA). The percentage of positive cells and the mean fluorescence intensity (arbitrary units) for a specific marker were calculated using FACSDiva software (BD Biosciences, San Jose, CA, USA). For purpose analysis 30000 events were recorded. 2.5. Evaluation of Immunological Mediators and Hormones Levels of TGF-��, IFN-��, and IL-6 were measured in plasma using commercially available high-sensitivity ELISA kits according to the instructions of http://www.selleck.co.jp/products/pembrolizumab.html the manufacturer. Detection limits were 0.07?pg/mL for IL-6 (R&D Systems, Inc. Minneapolis, USA), 125?pg/mL for TGF-��, and 4.7?pg/mL for IFN-�� (both from BD Bioscience, San Jose, CA, USA). Cortisol and DHEA-sulphated (DHEA-S) serum concentrations were assessed by electrochemiluminescence (Cobas e411, Roche, Germany). 2.6. Statistical Analysis Comparisons between groups (TB versus HCo) were performed by nonparametric methods, (Kruskall-Wallis analysis of variance and Mann-Whitney U test). Correlations between variables were assessed by Spearman's rank test. Data were considered statistically significant when P selleck chemical There were no differences with respect to age, sex distribution, BCG-vaccination, and peripheral CD8+ T cells frequency. However, we found that TB patients presented a significant diminution in CD4+ T cell levels (P