Legend max mouse ifnγ or il 17a elisa kits

96-well plates were incubated with or without 100 µl of diluted anti-TCRγδ antibody (BioXCell; clone: UC7-13D5; concentration: 4 µg/ml) at 37 °C and 5% CO₂ for 4 h to bind the antibody to the plate and washed twice with PBS to remove unbound antibody. Spleens were harvested from either uninfected C57BL/6, TCRβ^−/−, or TLR2^−/− mice, lysed of RBCs, washed, and processed for single-cell suspensions through a 70 μm pore filter. These cells were counted and either (i) 10⁵ live splenocytes in 100 µl complete RPMI were seeded into a 96-well plate with plate-bound anti-TCRγδ antibody or (ii) 10⁵ live γδ T cells were purified in 100 µl complete RPMI via a mouse γδ T cell isolation MACS separation kit (Miltenyi Biotec; catalog number 130-092-125) using an LD and MS column according to the manufacturer’s directions. 10⁵ live or HK C. neoformans WT, C. neoformans Δsgl1, C. neoformans Δcap59Δsgl1, or sterile PBS in 100 µl of complete RPMI were added to the appropriate wells in a 1:1 ratio to the γδ T cells. Seeded plates were incubated at 37 °C and 5% CO₂ for 5 days. Supernatants were collected 1, 3, and 5 days post incubation and stored at −80 °C until the ELISA was performed. The supernatants were divided and examined for IFNγ and IL-17A production using Legend Max Mouse IFNγ or IL-17A ELISA kits (BioLegend) following the manufacturer’s instructions exactly.