Pe conjugated anti human cd25

The eBioscience™ Cell Stimulation Cocktail (lot no.: 2046941) was purchased from Thermo Fisher Scientific Inc. (USA). FITC-conjugated anti-human CD4 (lot no.: B262223), PerCP-Cy5.5-conjugated anti-human CD3 (lot no.: B560835), Brilliant Violet 510-conjugated anti-human IFN-γ (lot no.: B323449), PE-conjugated anti-human IL-17A (lot no.: B315520), APC anti-human FOXP3 (lot no.: 1995356), PE-conjugated anti-human CD25 (lot no.: 2173867), Brilliant Violet 421-conjugated anti-human CD274 (lot no.: B320732), PE/Cyanine7-conjugated anti-human CD279 (lot no.: B318888), Fixation/Permeabilization Concentrate (lot no.: 2084746), eBioscience™ Fixation/Perm Diluent (lot no.: 2047346), Permeabilization Buffer 10× (lot no.: 2060497), and FIX&PERM (Fixation Medium A, Permeabilization Medium B) (lot no.: 17159) were purchased from Biolegend (USA).

In order to determine the activation of CD4⁺ T cells, PBMCs were incubated with FITC-conjugated anti-human CD4, APC-conjugated anti-human CD44, PE-conjugated anti-human CD25, PE-cy7-conjugated anti-human CD69 or appropriate isotypes (eBioscience, San Diego, CA) for 30 minutes at 4°C. FACScan flow cytometer (BD Biosciences, San Diego, CA) and FlowJo software (Tree Star, Inc. Ashland, USA) were used for flow cytometry analysis.