THP-1, or immortalized WT or NALP3−/− macrophages were attached to 35 mm 6-well plates. Cells were then transfected with siRNA oligonucleotides (50 nM) for 24 h using Lipofectamine 2000 in 2 ml of SF OPTI-MEM® I. The following day, cells were washed with SF OPTI-MEM® I and then transfected with poly (dA:dT) (1 µg/ml) for 6 hours with Lipofectamine 2000 in 2 ml of SF OPTI-MEM® I. The culture supernatants and cells were separated and then processed for immunoblot analysis with anti-caspase-1 and anti-IL-1β specific antibodies.
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