Figure 3.

Generation of bispecific ABA against TcdA and TcdB. A, Diagram of the ABA fusion protein. AH3/E3/E3/AA6 heavy-chain-only antibody (V_HH) subunits were separated by a flexible linker sequence (FS). A His(6)-tag and E-tag were genetically fused at the N-terminal and C-terminal, respectively, of the ABA molecule. B, Checkerboard Enzyme-linked immunosorbent assay analysis of ABA competition with individual components. Serially diluted ABA (ng/mL) was mixed with different serially diluted thioredoxin/V_HH fusion proteins targeting TcdA (AH3, AA6, or AC1) or TcdB (E3 or B12) and added to microplates coated with TcdA and TcdB, respectively. Biotinylated anti-thioredoxin V_HH was used for detection.