Ab9563

In vivo experiments were performed using age and sex-matched WT, Tnfr1^−/−, and Cd4^−/− C57BL/6J mice (Jackson Laboratories). Mice were anaesthetized with 100 mg/kg ketamine and 5 mg/kg xylazine, infected intranasally with MRSA, S. aureus 502A, K. pneumoniae ST 258 or P. aeruginosa PAK (10⁷ cfu in 50 QL of PBS), and sacrificed 18-24 hours after infection. To deplete macrophages, 75 QL of clodronate liposomes or PBS liposome controls were inoculated intranasally 24 hours prior to infection with MRSA as previously described^{12 (link)}. For the neutralization of IL-16, 100 μg of anti-IL-16 antibody (ab9563; Abcam) or IgG isotype (Jackson ImmunoResearch) was injected intraperitoneally 2 hours prior to infection as previously described^{45 (link)}. Animal experiments were performed in accordance with the guidelines of the IACUC at Columbia University (protocol number AAAE5252 and AAAD0624).

Antibodies used in vitro were anti-EGFR (1005; Santa Cruz), anti-FasL (NOK1; BD Biosciences), anti-TLR2 (H 175; Santa Cruz), anti-TNFR1 (H-271; Santa Cruz) and anti-IL-16 (ab9563; Abcam). Respective IgG isotype controls were used for comparison; rabbit (2027; Santa Cruz) or mouse (2025; Santa Cruz). Inhibitors used were pancaspase inhibitor ZVAD-FMK (Santa Cruz), caspase-3 inhibitor (Calbiochem), caspase 1 inhibitor (Calbiochem), calpeptin (Calbiochem), BAPTA (Calbiochem) and EDTA (Sigma).