Anti cd3 apc 17 0032 82

Mesenteric lymph nodes were collected, ground and filtered through 100 μm cell strainers. Cells were counted and stimulated using Cell stimulation Cocktail plus transport inhibitors 500× (eBioscience) for 4 h. At the end of stimulation, cells were harvested, fixed using IC fixation buffer and permeabilized using 1× permeabilization buffer (eBioscience). Cells were divided into two parts then stained with antibodies as follows: (a) anti-CD3-APC (17-0032-82, eBioscience), anti-CD4-FITC (MA5-17443, eBioscience), and IL-17A-PerCP-Cyanine5.5 (45-7177-82, eBioscience); (b) anti-CD3-APC (17-0032-82, eBioscience), anti-CD4-FITC (MA5-17443, eBioscience), anti-IL-4-PE (12-7041-82, eBioscience) and anti-IFN gama-PerCP-Cyanine5.5 (45-7311-82, eBioscience). Single cell suspensions were examined using BD Accuri C6 and the data were analyzed using C6 software. The number of IL-17A⁺CD4⁺CD3⁺, IFNγ⁺ CD4⁺CD3⁺ and IL-4⁺ CD4⁺CD3⁺cells were calculated.