Figure 5.

Western blot analysis to probe for the expression of sdAbs in AE1 and AE5. Western blots were carried out using homogenized midgut tissues from 16 h post-bloodmeal WT, AE1, and AE5 mosquitoes (n = 30/group). Purified CC3 and CA6 sdAbs (Liu et al., 2019) were used as controls and all samples were probed with AffiniPure Goat Anti-Alpaca IgG, VHH domain primary antibody. The presence of a 15 kDa band confirms the expression of the individual, cleaved sdAb proteins (CA6 and CC3) in transgenic, but not in WT, mosquito midgut samples. The presence of a 30 kDa band in the transgenic samples may indicate unsuccessful ribosomal skipping of the P2A linker sequence located between the CA6 and CC3 sdAbs, resulting in a (uncleaved) CA6-CC3 fusion protein.