Cell-Based TLR Activation Assays.

Various CEP adducts and TLR agonists were tested in HEK293 or THP-1 cells. Readouts were NFkB reporter signal or IL-8 secretion (columns), as indicated. In addition, the same wells were analyzed for viability with the CellTiter-Glo kit (axis on right; square symbols) to ensure that any lack of activation was not due to cell toxicity. HEK293 cells were treated with the following reagents: HSA-CTL1, HSA-CTL2, or HSA-CEP: 0, 3.9, 7.8, 15.6, 32.6, 62.5, and 125 and 250 µg/ml; Pam3CSK4∶ 0, 1.5, 3.2, 6.3, 12.5, 25, 50, 100 ng/mL. THP-1 cells were treated with the following reagents: HSA-CTL1, HSA-CTL2, or HSA-CEP: 62.5, 125, and 250 µg/mL; Pam3CSK4∶ 4, 20, and 100 ng/mL; FSL-1∶0.4, 2, and 10 ng/mL; LPS: 4, 20, and 100 ng/mL; R837 or R848∶0.4, 2, and 10 µM; ODN2006G5∶0.2, 1, and 5 µM.