466680 Silkworm Expression System Suitable for High-Throughput Screening of Antibody Fragment
We attempted to produce single chain fragments of variable region (scFv), where heavy chain of variable region (VH) and light chain of variable region (VL) are linked via a peptide linker. First, we inserted the scFv gene from the antibody with affinity for CD3 on lymphocyte cell and with affinity for epidermal growth factor receptor (EGFR) on cancer cells into baculovirus vectors and we transduced silkworm pupae with the vectors. The pupas were smashed and then ultracentrifuged. From the supernatant after ultracentrifugation, the scFv were purified by means of immobilized metal affinity chromatography and size exclusion chromatography. Finally, we measured the binding affinity of scFvs for each target molecule by means of flow cytometry and surface plasmon resonance.
In this study, 3 and 8 kinds of scFvs were prepared from anti-CD3 and anti-EGFR antibody, respectively. The scFvs were purified from the supernatant after ultracentrifugation, although the expressed amounts were varied. We confirmed that 8 out of 11 scFvs bound to each target molecule. From these results, we concluded that scFvs can be prepared from silkworms.