- 著者
-
LIU Xiaobo
EICHENBERGER Michael
FUJIOKA Yuuichiro
DONG Jinhua
UEDA Hiroshi
- 出版者
- The Japan Society for Analytical Chemistry
- 雑誌
- Analytical Sciences (ISSN:09106340)
- 巻号頁・発行日
- vol.28, no.9, pp.861-867, 2012
- 被引用文献数
-
10
The development of a rapid and specific assay for 17β-estradiol (E2) will accelerate its <i>in vitro</i> 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<sub>H</sub> and V<sub>L</sub>) dissociated in the absence of an antigen. The V<sub>H</sub> 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<sub>H</sub>, 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.