Construction and panning of single chain Fv phage display library against anti-DR5

The study aim to construct single chain Fv(scFv) library against human death receptor 5 by phage display technique and to obtain the anti-DR5 scFv clones selected from the library.Total RNA was extracted from splenocytes of the Balb/c mice immunized with recombinant human DR5.Complementary DNA fragments of variable heavy(VH) and variable light(VL) chains of antibodies were prepared by RT-PCR and assembled into scFv.Then the scFv fragment was inserted into the phagemid vector PAK100 through SfiⅠsites.The phagemids containing scFv cDNA were transformed into E.coli XL1-Blue bacterial cells to generate scFv antibody library with a diversity of approximately 1×106.Five rounds of enrichment and panning were performed to select specific anti-DR5 scFv from the library.The antigen binding activity was evaluated via ELISA assay.Sequencing analysis of one positive clone showed that the anti-DR5 scFv was 747 bp,and coded 249 amino acids.The results mean an anti-DR5 scFv is obtained from scFv antibody library,which set a base for seeking novel antibodies of using in immunotherapy.