To determine the associations between rs1800629 genotypes and TNF-a levels in PBMCs, we investigated 15 subjects with rs1800629GG genotype, 8 subjects with GA genotype and 1 subject with AA genotype. PBMCs were derived by using Ficoll gradient density centrifugation method. Isolated PBMCs were plated at a density of 1610 6 cells/ml in 24-well plates and cultured in RPMI 1640 medium with 10% FBS at 37 uC with 5% CO 2 . The cells were then incubated for 6 hours in presence or absence of 100 ng/ml Escherichia coli 0111:B4 LPS (Sigma, USA). After incubation, supernatants were harvested and stored at 280uC until use.