A phage-displayed single chain variable fragment that interacts with hepatitis B core antigen: library construction, selection and diagnosis.
BACKGROUND
Phage display is an alternative method for constructing and selecting antibodies with desired specificity towards an antigen.


OBJECTIVE
To construct a library of single chain variable fragment (ScFv) towards hepatitis B core antigen (HBcAg). To isolate a ScFv phage clone that interacts with HBcAg and to develop a phage-ELISA for detecting the antigen.


METHODS
Mice were inoculated with HBcAg and RNA was extracted from their spleen cells. The genes encoding heavy (V(H)) and light (V(L)) chains were amplified, linked via PCR and cloned into a phagemid vector. Phage particles displaying ScFv were panned against HBcAg and a selected clone was characterized and employed as a diagnostic reagent for detecting HBcAg in serum samples.


RESULTS
A phage clone that interacts with HBcAg was selected from the antibody library. The binding of the phage to HBcAg was inhibited by a cyclic peptide bearing the WSFFSNI sequence. A phage-ELISA was established using the recombinant phage and as low as 10ng of HBcAg can be detected by the assay.


CONCLUSIONS
The ScFv displayed on the surface of filamentous phage is an alternative choice for diagnosis of HBcAg in serum samples.
Tan
Geok Hun
GH
Department of Microbiology, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, 43400 UPM Serdang, Selangor, Malaysia.
Yusoff
Khatijah
K
Seow
Heng Fong
HF
Tan
Wen Siang
WS
D000595:Amino Acid Sequence; D000818:Animals; D000914:Antibodies, Viral / Q000235:genetics* / Q000276:immunology*; D000918:Antibody Specificity; D001483:Base Sequence; D005260:Female; D006509:Hepatitis B / Q000175:diagnosis / Q000276:immunology*; D006512:Hepatitis B Core Antigens / Q000032:analysis / Q000235:genetics* / Q000276:immunology; D007143:Immunoglobulin Heavy Chains / Q000235:genetics* / Q000276:immunology; D007147:Immunoglobulin Light Chains / Q000235:genetics* / Q000276:immunology; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D019151:Peptide Library
D000914:Antibodies, Viral; D006512:Hepatitis B Core Antigens; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains; D019151:Peptide Library