title: [Expression, purification and activity analysis of anti-human transferrin receptor scFv].
abstract: Human transferrin receptor (TfR) was isolated from homogenates of placental tissues by affinity chromatography on transferrin-Sepharose, and then used to screen human scFv against it from a fully-synthesized phage scFv library. After verifying the specificity, gene fragment of one of the selected scFv was inserted into the plasmid pET22b(+) and transformed into E. coli BL21(DE3) . Expression of scFv in transformant was induced with 0.5mmol/L IPTG. ELISA assay on HeLa cells showed that scFv protein could recognize and bind to TfR on the surface of HeLa cells. The scFv was purified by one-step affinity chromatography with Ni+ -NTA agarose, and injected into Kunming mouse via tail veins. This scFv was detected in brain tissues 1h later by capillary depletion method, which indicates that scFv protein can permeate through the blood brain barrier by mediation of the TfR receptor. Our works lay the foundation for the treatment of tumors and central nervous system diseases.
authors.affiliation: Beijing Institute of Biotechnology, Beijing 100071, China.
authors.forename: Li-Xia
authors.initials: LX
authors.lastname: Zhao
authors.forename: Bing
authors.initials: B
authors.lastname: Yan
authors.forename: Long
authors.initials: L
authors.lastname: Xu
authors.forename: Shi-Wei
authors.initials: SW
authors.lastname: Jiang
authors.forename: Ying-Ying
authors.initials: YY
authors.lastname: Zhang
authors.forename: Zhi-Xin
authors.initials: ZX
authors.lastname: Yang
authors.forename: Xiao-Wei
authors.initials: XW
authors.lastname: Zhou
authors.forename: Pei-Tang
authors.initials: PT
authors.lastname: Huang
mesh_terms: D000818:Animals; D000888:Antibodies, Anti-Idiotypic / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D007128:Immunoglobulin Fragments / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D007135:Immunoglobulin Variable Region / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D051379:Mice; D011990:Receptors, Transferrin / Q000235:genetics / Q000276:immunology*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D014168:Transferrin / Q000378:metabolism
chemical_list: D000888:Antibodies, Anti-Idiotypic; D007128:Immunoglobulin Fragments; D007135:Immunoglobulin Variable Region; D011990:Receptors, Transferrin; D011994:Recombinant Proteins; D014168:Transferrin