[Construction of human phage-displayed scFv library and selection of the ScFv against rabies virus].
OBJECTIVE
To construct a human phage displayed single chain Fv antibody (scFv) library and screen specific scFv against rabies virus.


METHODS
Using a phage-display technique, IgVH and IgVL genes were cloned from peripheral blood lymphocytes from three donors immunized with the WISTAR PM strain vaccine. Genes encoding scFv fragments were constructed by random linkage of VH gene and VL gene by SOE PCR, and then the constructed scFv genes were cloned into phagemid vector and transfected into E. coli TG1. The recombinant phages were screened by four rounds of panning with rabies virus vaccine. The positive recombinant phages were sequenced and antigen-binding activity of recombinant scFv was detected by competition ELISA.


RESULTS
A human phage-displayed scFv library with about 7 x 10(8) sink size was constructed. A new human scFv was selected, which bound specifically to rabies virus and had higher affinity.


CONCLUSIONS
The results lay a solid foundation for preparation of human engineering antibody to rabies virus reported herein with higher affinity.
Zhao
Xiao-ling
XL
Institute of Microbiology and Epidemiology, the Academy of Military Medical Sciences, Beijing 100071, China. xlzhao985@sohu.com
Yin
Jun
J
Wang
Hui
H
Jiang
Ming
M
Hou
Xiao-jun
XJ
D000595:Amino Acid Sequence; D000914:Antibodies, Viral / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D000918:Antibody Specificity; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007128:Immunoglobulin Fragments / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D007143:Immunoglobulin Heavy Chains / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D007147:Immunoglobulin Light Chains / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D007135:Immunoglobulin Variable Region / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D008969:Molecular Sequence Data; D019151:Peptide Library; D011820:Rabies virus / Q000276:immunology*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D020539:Sequence Analysis, Protein; D012995:Solubility
D000914:Antibodies, Viral; D007128:Immunoglobulin Fragments; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains; D007135:Immunoglobulin Variable Region; D019151:Peptide Library; D011994:Recombinant Proteins; C055966:immunoglobulin Fv