Ultra Q-bodies: Quench-based Antibody Probes That Utilize Dye-dye Interactions with Enhanced Antigen-dependent Fluorescence

Overview

Journal Sci Rep

Specialty Science

Date 2014 Apr 12

PMID 24721819

Citations 18

Authors

Ryoji Abe

Hee-Jin Jeong

Dai Arakawa

Jinhua Dong

Hiroyuki Ohashi

Rena Kaigome

Fujio Saiki

Kyosuke Yamane

Hiroaki Takagi

Hiroshi Ueda

Affiliations

Soon will be listed here.

Abstract

Recently, we described a novel reagentless fluorescent biosensor strategy named Quenchbody, which functions via the antigen-dependent removal of the quenching effect on a fluorophore that is attached to a single-chain antibody variable region. To explore the practical utility of Quenchbodies, we prepared antibody Fab fragments that were fluorolabeled at either one or two of the N-terminal regions, using a cell-free translation-mediated position-specific protein labeling system. Unexpectedly, the Fab fragment labeled at the heavy chain N-terminal region demonstrated a deeper quenching and antigen-dependent release compared to that observed using scFv. Moreover, when the Fab was fluorolabeled at the two N-termini with either the same dye or with two different dyes, an improved response due to enhanced quenching via dye-dye interactions was observed. On the basis of this approach, several targets, including peptides, proteins, and haptens, as well as narcotics, were quantified with a higher response up to 50-fold. In addition, differentiation of osteosarcoma to osteoblasts was successfully imaged using a similarly fluorolabeled recombinant Fab protein prepared from E. coli. Due to its versatility, this "Ultra-Quenchbody" is expected to exhibit a range of applications from in vitro diagnostics to the live imaging of various targets in situ.

Citing Articles

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