Briefly, Raw 264.7 cells were seeded on a
96-well plate at a density of 105 cells/well in complete DMEM
and incubated at 37 C overnight. The cells were pre-treated with
samples in fresh DMEM without FBS for 1 h followed by treatment
with 50 ng/mL of lipopolysaccharide (LPS) and 10 ng/mL of
interferon gamma (IFN-c) for another 24 h to induce NO. Cell
culture supernatant (100 lL) was reacted with 100 lL of Griess
reagent to form pink color which was measured at 540 nm. Sodium
nitrite was used as the standard.