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. 2022 May 23;10(5):828. doi: 10.3390/vaccines10050828

Figure 1.

Figure 1

IL-1β antigen design. (a) Graphical representation of full-length (FL) murine IL-1β and the five IL-1β antigen constructs. FL murine IL-1β consists of a pro-peptide (aa1-117) that is cleaved to release the mature active 17kDa IL-1β protein (blue). The muIL-1β antigens were N-terminally fused to a 6xHis-tag (red), followed by a TEV site (gray). A split-protein binding Tag (green) was added to the C-terminus of the antigen, separated by a flexible linker. Mutations (yellow) were introduced into the WT sequence to reduce the affinity of the antigens for IL-1R1. (b) A HEK293 cell assay was used to assess the biological activity of the produced protein antigens. A dilution series of the IL-1β proteins (3-fold, starting from 100 ng/mL) was added to engineered HEK293 cells, and the biological activity was estimated based on SEAP detection (620 nm) after overnight incubation. Tested antigens include IL-1β-Tag (blue, EC50 = 0.75), IL-1β(R11G)-Tag (magenta, EC50 = 2.7), IL-1β(Q15G)-Tag (orange, EC50 = 5.8), IL-1β(H30G)-Tag (grey, EC50 = 2.4), IL-1β(N32G)-Tag (red, EC50 = 0.89). Positive controls include, muIL-1β (black, EC50 = 0.21) and huIL-1β (cyan, EC50 = 0.25). A technical replicate of this assay showed the same tendency (Figure S1A). The EC50 values are shown in Figure S1B.