title: An hepatitis B virus surface antigen specific single chain of variable fragment derived from a natural immune antigen binding fragment phage display library is specifically internalized by HepG2.2.15 cells.
abstract: Hepatitis B virus surface antigen (HBsAg), a specific antigen on the membrane of hepatitis B virus (HBV)-infected cells, provides a perfect target for therapeutic drugs. In order to mediate successful targeted delivery of these therapies, it is essential to have antibodies that recognize HBsAg with high specificity and affinity. In this report, we constructed a natural immune antigen binding fragments (Fab) antibody phage display library against HBsAg and after three rounds of panning, five Fab fragments with significant HBsAg binding ability were selected and analysed. DNA sequencing revealed that all the light chains had the same sequence, while all the Fd genes exhibited different sequences. For further application, all of the Fab antibodies were reconstructed into single chain antibodies (scFvs) and expressed in Escherichia coli BL21 cells. Indirect enzyme-linked immunosorbent assay analysis demonstrated that all five scFvs maintained a high affinity for HBsAg and could bind HBsAg on the membrane of HBV-infected cells. Indirect fluorescent staining analysis revealed that one of the scFvs (scFv15) could be internalized into HBsAg-positive HepG2.2.15 cells through clathrin-mediated endocytosis pathway. The internalizing scFv15 antibody would have great potential for the targeted delivery of therapeutics to HBV-infected cells.
authors.affiliation: State Key Laboratory of Cancer Biology, Department of Immunology, Fourth Military Medical University, Xi'an, China.
authors.forename: W-H
authors.initials: WH
authors.lastname: Wen
authors.forename: W-J
authors.initials: WJ
authors.lastname: Qin
authors.forename: H
authors.initials: H
authors.lastname: Gao
authors.forename: J
authors.initials: J
authors.lastname: Zhao
authors.forename: L-T
authors.initials: LT
authors.lastname: Jia
authors.forename: Q-H
authors.initials: QH
authors.lastname: Liao
authors.forename: Y-L
authors.initials: YL
authors.lastname: Meng
authors.forename: B-Q
authors.initials: BQ
authors.lastname: Jin
authors.forename: L-B
authors.initials: LB
authors.lastname: Yao
authors.forename: S-Y
authors.initials: SY
authors.lastname: Chen
authors.forename: A-G
authors.initials: AG
authors.lastname: Yang
mesh_terms: D000915:Antibody Affinity; D000918:Antibody Specificity; D001402:B-Lymphocytes; D002460:Cell Line; D006509:Hepatitis B / Q000517:prevention & control; D006510:Hepatitis B Antibodies / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D006514:Hepatitis B Surface Antigens / Q000378:metabolism*; D017325:Hepatitis B Vaccines / Q000008:administration & dosage / Q000276:immunology; D006515:Hepatitis B virus / Q000276:immunology* / Q000378:metabolism; D006801:Humans; D007135:Immunoglobulin Variable Region / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D008969:Molecular Sequence Data; D019151:Peptide Library
chemical_list: D006510:Hepatitis B Antibodies; D006514:Hepatitis B Surface Antigens; D017325:Hepatitis B Vaccines; D007135:Immunoglobulin Variable Region; D019151:Peptide Library