Enzyme-Linked Immunosorbent Assay (ELISA)
Cell supernatants were centrifuged at 450×g for 5 mins and stored at -20°C until usage. Cytokine concentrations were determined using ELISA Duoset kits for mouse IL-1β, IL-6 and TNF-α according to the manufacturer’s protocol (#DY401, #DY406 and #DY410, respectively; R&D System, Minneapolis, Canada). Absorbance at the wavelength of 450 nm was measured using a plate reader (SpectraMax 190; Molecular Devices, San Jose, California, USA) as a correction wavelength of 540 nm. Concentrations were calculated using eight-parameter fit curve.