Mouse interleukin 36γ elisa kit

For Oil Red O (A12989.14 Thermo Fisher Scientific) staining, CD11c⁺ or CD11c⁻ macrophages were fixed in 4% paraformaldehyde for 15 min, washed twice with dH₂O, permeabilized with 60% isopropanol (190764, Sigma for 5 min, stained with an Oil Red O (Stock solution 75 mg in 25 mL 100% isopropanol and working solution 3 parts Oil Red O to 2 parts dH₂O 0.2 μm filtered) for 20 min, images were collected using an Echo RVL2-K microscope.
For FA treatment effects, CD11c⁺ or CD11c⁻ macrophages were treated with 200 μM PA, DPA or control BSA in the presence or absence of 1 mM N-acetyl-L-cysteine (NAC, A7250, Sigma-Aldrich) or 5μM Diphenyleneiodonium chloride (DPI, Sigma-Aldrich) for 24 h for gene expression analysis by real-time PCR. For IL-36 intracellular staining, cells were treated with 200 μM PA, DPA or control BSA for 48 h. After surface staining with anti-mouse CD11c and anti-mouse F4/80, cells were permeabilized for intracellular staining of IL-36α (32103-05161, Assaypro) or IL-36γ (PAL621Mu01, Cloud-clone Corp). For measurement of IL-36γ concentration in cultural supernatants, CD11c⁺ macrophages were treated with 200 μM PA, DPA or control BSA for 48 h. Supernatants were collected and detected using mouse Interleukin-36γ ELISA Kit (MBS288173, Mybiosource) according to the manufacturer’s protocol.