Anti-human antibodies for TLR 2, 3, 4 and 9 (eBioscience, USA) and TLR 7 and TLR 8 (Imgenex, USA) were used for the staining. For surface staining of TLR 2 and 4, 100 ml of whole blood was lysed with BD FACS lysis solution (BD Biosciences, USA), washed, fixed and processed for staining with anti-human TLR 2 (FITC conjugated) and TLR 4 (PE conjugated) antibodies respectively. For intracellular staining, 100 ml of the whole blood was fully lysed with BD FACS lysing solution (Becton Dickinson), washed twice with Perm-wash buffer (BD bioscience) and processed for staining with the following anti-human antibodies: PE-TLR 3, FITC-TLR 7, FITC-TLR8 and PE-TLR 9 respectively. Stained cells were resuspended in 500 ml 1% paraformaldehyde, analysed on FACScalibur flow cytometer (Becton Dickinson).On the basis of forward and side scatter plot lymphocytes and monocytes were gated and data analysis was done using BD FACSDivasoftware.TLR levels were expressed as median fluorescence intensity (MFI).