Figure 1

The purified sporozoites and their TLRs activities.

(A), Microscopy of the contamination in the sporozoites before or after purification by DEAE-cellulose chromatography. (B), After HEK 293FT cells/well were plated in 96-well plate for 24 h, each well was transfected with the indicated TLRs along with TK-RL and pBIIx-luc or pGL3-IFN-β. 24 h later, cells were stimulated with the indicated positive TLR agonist, SPZ lysate or NSG lysate. After incubation for 6 h or 18 h, either NF-κB (TLR2, TLR4, TLR5, TLR7 and TLR9) or IFN-β gene (TLR3) reporter activity was determined. Both NF-κB and IFN-β gene reporter activity were expressed as the ratio of Renilla luciferase activity to firefly luciferase activity. (C), TLR2-transfected HEK293FT cells were stimulated with whole sporozoites lack of infectivity (cell/sporozoite = 1:5) for 6 h, then the NF-κB gene reporter activity was then determined. (D), Peritoneal macrophages were stimulated with LPS, SPZ or NSG treated with or without PmB and the concentrations of both TNF-α and IL-6 in the supernatants were determined using CBA beads. All the experiments were repeated three times and all data were presented as the mean ± SD, *p < 0.05; **p < 0.01.