2.6. Nitricoxide(NO)productionandev

2.6. Nitricoxide(NO)productionandevaluationofanti-allergiceffects
Viable and heat-treated (100 C, 5 min) bacterial cells were added at concentrations of 1 107 colony-forming units (CFUs)/ well with or without lipopolysaccharide (LPS) in RAW 264.7 cells. After 3 h of incubation, the conditioned media (100 mL) were allowed to react with an equal volume of Griess reagent (Sigma) for 15 min at room temperature. The optical density was determined at 540 nm using an ELISA plate reader (Molecular Devices, Sunnyvale, CA, U.S.A.). NO production was evaluated by comparison of the optical density with the standard curve obtained with sodium ni- trite (SigmaeAldrich). Viableandheat-treated(100 C,5min)bacterialcellswereadded at concentrations of 1 107 CFUs/well with or without LPS in RAW 264.7 cells for analysis of cytokine. After 3 h of incubation,