Procartaplex porcine cytokine chemokine panel 1

The supernatants of SF were diluted 1:4 in PBS and stored at −20 °C. These supernatants were thawed and IFNα, IFNγ, IL-1b, IL-4, IL-6, IL-8, IL-10, IL-12p40 and TNFα were analyzed using a multiplexed immunoassay. The measurements were performed according to the manufacturer’s instructions by Luminex xMAP technology using the ProcartaPlex Porcine Cytokine & Chemokine Panel 1 (eBioscience, San Diego, CA, USA; catalog number EPX090–60829-901). The concentrations of the different cytokines were expressed as pg/ml, and calculated according to a standard curve.

The pericardial fluid supernatants and plasma samples were stored at -80°C until further processing. Cytokines levels of IFNα, IFNγ, IL-1b, IL-10, IL-12p40, IL-4, IL-6, IL-8 and TNFα were analyzed using the Luminex xMAP technology, a multiplexed sandwich immunoassay. The measurements were determined using the ProcartaPlex Porcine Cytokine & Chemokine Panel 1 (catalog number EPX090-60829-901) according to the manufacturer’s instructions (eBioscience). The concentrations of the different cytokines were expressed as pg/mL, and calculated according to a standard curve.

PAMs were seeded in 24-well plates (10⁶ cells/well) in RPMI 1640 medium supplemented with 10% FBS, 100 IU/mL of penicillin/streptomycin, 2 mM L-glutamine, and 0.5% nystatin (sigma-MERCK) and incubated overnight at 37 °C in 5% CO₂. Afterwards, the medium was removed and replenished with fresh RPMI either with or without 5 µg/mL of GFP POMs. After a 24 h incubation at 37 °C, 5% CO₂, cell supernatants were collected and cytokines were determined using the Luminex xMAP technology and the ProcartaPlex Porcine Cytokine & Chemokine Panel 1 (Thermo Fisher Scientific). Cytokine concentrations were calculated using the xPONENT 4.3 software (Luminex, Austin, TX, USA) and expressed as pg/mL (except for TNF, for which a technical problem with the standard curve occurred and only the mean fluorescence intensity (MFI) values could be assessed).