Il 12p70

U-Plex 10 Assay Kits for analysis of IL-1b, IL-6, IL-10, IL-12p70, TNF-α, VEGFa, IP10, MCP1, TARC, and MDC were purchased from Meso Scale Diagnostics (MSD, USA). The assay was performed as described by the manufacturer. ELISA kits to measure MMP1 (DY901B), MIP1-a (DY270), MIP1-b (DY271), and IL-2 (DY202) were obtained from R&D Systems. The supernatants were tested according to the manufacturer's instructions in 96-well half area plates using 50 μL sample/test.

The BALF supernatant samples were analysed using MSD pro-inflammatory panel V-Plex including the following analytes: IFN-γ, IL-10, IL-12p70, IL-1β, IL-2, IL-4, IL-5, IL-6, KC/GRO and TNF-α (Meso Scale Diagnostics, Rockville, MD). The analysis was performed according to the manufacturer’s instruction.

Rabbit anti-human phospho p65 NF-κB, PathScan^® Signaling Nodes Multi-Target Sandwich ELISA, PathScan^® signaling NF-κB ELISA, and PathScan^® Sandwich ELISA Lysis Buffer were obtained from Cell Signaling Technology (Pickering, ON, Canada). Cy3-conjugated goat anti-mouse IgG was purchased from Invitrogen Canada (Burlington, ON, Canada). Bordet-Gengou blood agar plates were from BD Biosciences (Mississauga, ON, Canada). Syto9 and Propidium Iodide were from ThermoFisher (Waltham, MA). Wild-type B. pertussis (Tohama I) was propagated at Sanofi Pasteur (Toronto, ON, Canada). Human lung carcinoma epithelial A549 cell line was obtained from ATCC (Old Town Manassas, VA). For the cytokine detection (Eotaxin, Eotaxin-3, IL-10, IL-12/IL-23p40, IL-12p70, IL-13, IL-15, IL-16, IL-17A, IL-1α, IL-1β, IL-2, IL-4, IL-5, IL-6, IL-7, IL-8, IL-8 (HA), IL-21, IL-22, IL-23, IL-27, IL-31, IP-10, MCP-1, MCP-4, MDC, MIP-1α, MIP-1β, MIP-3α, TARC, TNF-α, TNF-β, VEGF-A, GM-CSF, and IFN-γ), V-PLEX Human Cytokine 36-Plex Kit (K15089D-2) was used for screening and only 20 selected cytokines were further tested on 20-plex cytokines (Meso Scale Discovery, Rockville, MD).