title: Characterization and enzyme-conjugation of a specific anti-L1 nanobody.
abstract: Persistent infection of the human papillomaviruses (HPV) has been shown to result in cervical cancer and intraepithelial neoplasia. Early detection and screening programs are essential strategies against cervical cancer. A nanobody is the smallest antigen-binding fragment known and is derived from a camelid heavy-chain antibody. This tiny protein shows high solubility and stability. It can be produced cost-effectively with high yield production. In this study, we enriched a nanobody library against the L1 protein of HPV. Several colons were selected from this enriched library using monoclonal phage-enzyme linked immunosorbent assay (phage-ELISA) and analyzed for identification of nanobody genes. The expression of nanobody fragments was performed in Rosetta gami2. The C74 nanobody that showed strong binding to the L1 protein of HPV16 was selected, purified, and characterized by Western blotting and ELISA. The selected nanobody was tested for sensitivity, specificity, and affinity. A nanobody conjugated to horseradish peroxidase (HRP) was selected and used for detection of L1 protein of HPV16. This study demonstrates that the C74-HRP, due to its specificity and good binding affinity for a specific viral antigen, is a potential diagnostic tool that can be used as a promising reagent for the new generation of HPV diagnosis approaches.
authors.affiliation: Department of Biology, Faculty of Sciences, University of Isfahan, Isfahan, Iran.
authors.forename: Sara
authors.initials: S
authors.lastname: Minaeian
authors.forename: Fatemeh
authors.initials: F
authors.lastname: Rahbarizadeh
authors.forename: Sayyed Hamid
authors.initials: SH
authors.lastname: Zarkesh Esfahani
authors.forename: Davoud
authors.initials: D
authors.lastname: Ahmadvand
mesh_terms: D000914:Antibodies, Viral / Q000737:chemistry / Q000276:immunology* / Q000378:metabolism*; D000918:Antibody Specificity; D000937:Antigen-Antibody Reactions / Q000276:immunology; D015153:Blotting, Western; D036022:Capsid Proteins / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay; D006735:Horseradish Peroxidase / Q000737:chemistry / Q000378:metabolism*; D009856:Oncogene Proteins, Viral / Q000276:immunology*; D057127:Single-Chain Antibodies / Q000737:chemistry / Q000276:immunology* / Q000378:metabolism*
chemical_list: D000914:Antibodies, Viral; D036022:Capsid Proteins; C114580:HPV L1 protein, Human papillomavirus; D009856:Oncogene Proteins, Viral; D057127:Single-Chain Antibodies; D006735:Horseradish Peroxidase