Expression of anti-RBC blood group A substance ScFv by using phage display technology

Objective:To construct a phage-displayed ScFv antibody of 50A hybridoma anti-blood group A substance.Methods:Based on recombinant phage display techniques,the construction,screening and expression of functional single-chain Fv fragment(ScFv) from murine hybridoma 50A which secrets antibody specifity binding to human blood group A substance was performed. By RT-PCR for VH and VL genes assembly of ScFv genes and cloning into phagemid pCANTAB5E, transformation of E.coli XL-Blue cells and rescuing with M13KO7 helper phage. The recombinant phages were panned by whole red blood cells over three rounds. Finally, the positive recombinant phages were sequenced and identified by immuno-blot assay. Results:A recombinant phage ScFv library with titer of 4×10 9 pfu/ml was established. The result of immuno-blot indicated that the phage-displayed 50A-ScFv retained the affinity and specificity of the original intact antibody to blood group A substance.Conclusion:Single chain antibody fragment 50A-ScFv displayed on the surface of filamentous phage was successfully produced,which would be potentially useful in constructing engineering antibody fragments as blood grouping reagents.