title: Improved detection sensitivity and selectivity attained by open-sandwich selection of an anti-estradiol antibody.
abstract: The development of a rapid and specific assay for 17β-estradiol (E2) will accelerate its in vitro diagnostics and/or environmental pollution control. Here, we employed an open-sandwich (OS) selection scheme to improve the sensitivity for E2 in an OS immunoassay, which is based on antigen-dependent stabilization of the antibody (Ab) variable region, Fv, where the two domains (V(H) and V(L)) dissociated in the absence of an antigen. The V(H) domain of a cloned anti-E2 antibody displayed on M13 phage was randomly mutated, and after three OS biopanning rounds, a mutant that showed higher sensitivity in OS-ELISA for E2 was identified. Interestingly, compared with the wild-type V(H), the cross-reactivity of the mutant was significantly decreased for the analogous steroid testosterone, both in OS and competitive ELISAs. This is the first report concerning selection for an anti-hapten Ab without using any hapten-carrier conjugates, and the method will be especially suitable for selecting Ab fragments that show better performance in hapten OS immunoassays.
authors.affiliation: Department of Bioengineering, School of Engineering, The University of Tokyo, Japan.
authors.forename: Xiaobo
authors.initials: X
authors.lastname: Liu
authors.forename: Michael
authors.initials: M
authors.lastname: Eichenberger
authors.forename: Yuuichiro
authors.initials: Y
authors.lastname: Fujioka
authors.forename: Jinhua
authors.initials: J
authors.lastname: Dong
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Ueda
mesh_terms: D000906:Antibodies / Q000737:chemistry*; D000918:Antibody Specificity; D000937:Antigen-Antibody Reactions; D004958:Estradiol / Q000032:analysis*; D007118:Immunoassay; D015394:Molecular Structure
chemical_list: D000906:Antibodies; D004958:Estradiol