A phage-displayed single chain variable fragment that interacts with hepatitis B core antigen: Library construction, selection and diagnosis
Received 13 April 2006; received in revised form 11 August 2006; accepted 19 September 2006. published online 03 November 2006.
Abstract
Background
Phage display is an alternative method for constructing and selecting antibodies with desired specificity towards an antigen.
Objectives
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.
Study design
Mice were inoculated with HBcAg and RNA was extracted from their spleen cells. The genes encoding heavy (VH) and light (VL) 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.
Conclusion
The ScFv displayed on the surface of filamentous phage is an alternative choice for diagnosis of HBcAg in serum samples.
aDepartment of Microbiology, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, 43400 UPM Serdang, Selangor, Malaysia
bInstitute of Bioscience, Universiti Putra Malaysia, 43400 Serdang, Selangor, Malaysia
cDepartment of Clinical Laboratory Science, Faculty of Medicine and Health Science, Universiti Putra Malaysia, 43400 Serdang, Selangor, Malaysia
Corresponding author. Department of Microbiology, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, 43400 UPM Serdang, Selangor, Malaysia. Tel.: +60 3 89466715; fax: +60 3 89430913.
ABSTRACT