Figure 1.

Schematic diagram for mutants’ generation, screening, and selection of selective with desirable affinity antibody using yeast surface display. Step-1 represents the general protocol to generate a quality mutagenesis library using error-prone PCR, and the subsequent packaging of the library into yeast cells. Step-2 represents the screening of specific target antibodies after induction of the surface-displayed scFv library and incubation with the biotinylated protein antigens. Step-3 represents for selection of a single clone yeast surface display antibody with low cross-reactivity to chelator. The fusion protein is tethered to the yeast cell wall via disulfide bridges between the Aga2p protein and the Aga1p protein (which is covalently attached to the cell wall). The fusion protein also contains c-myc affinity tag (EQKLISEEDL) at the C-terminus can be used to monitor the full-length expression of the gene on the yeast surface by flow cytometry. Yeast displays functional antibodies that bind ligand can be identified and isolated by differential fluorescent staining of the antibodies and ligand.