Ifn γ clone 25723.11 tnf α clone mab11 perforin clone δg9

Intracellular cytokine staining and perforin assays were performed in samples from eight acute patients, six convalescent patients, and eight healthy controls, as described previously (Fu et al., 2010 (link); Das and Tripathy, 2014 (link)). Following co-incubation (effector/target) for 6 h, cells were stained with anti-human CD56 (clone B159) and CD3 (clone SK7) monoclonal antibodies (BD Biosciences) to enumerate the respective cell surface antigens. The CD3 and CD56 surface-stained cells were permeabilized for intracellular staining. The monoclonal antibodies, i.e., IFN-γ (clone 25723.11)/TNF-α (clone MAb11)/ perforin (clone δG9) (BD Biosciences) were added to the permeabilized cells, incubated, washed in BD Perm/Wash buffer (BD Biosciences) and fixed in paraformaldehyde. Cells were gated on NK and NKT-like cells. During acquisition, 5000 events were acquired within the lymphocyte gate.
Acquisition and analysis of the samples was performed in BD FACS ARIA II using the FACS DIVA software (BD Biosciences).

Intracellular cytokine staining and perforin assays were performed as described previously [30 (link), 33 (link), 35 (link)]. In vitro stimulation of effector cells with target cells and the incubation period were the same as described above. After incubation, cells were washed and stained with antibodies against CD56 (clone B159) and CD3 (clone SK7) (BD Biosciences, CA, USA). The surface-stained cells for NK/NKT-like cells were permeabilized using BD Cytofix/Cytoperm solution (BD Bioscience, CA, USA) and then subject to intracellular cytokine staining using IFN-γ (clone 25723.11)/TNF-α (clone MAb11)/perforin (clone δG9) (BD Bioscience, CA, USA) antibodies.
Acquisition and analysis of the samples were performed in the BD FACS ARIA II using FACS DIVA software (BD Biosciences, CA, USA).