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Sensors 2014, 14(7), 13285-13297; doi:10.3390/s140713285
Article

Strategy for Making a Superior Quenchbody to Proteins: Effect of the Fluorophore Position

 and *
Received: 27 May 2014; in revised form: 3 July 2014 / Accepted: 16 July 2014 / Published: 23 July 2014
(This article belongs to the Special Issue Immunosensors 2014)
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Abstract: Antibody-based sensors have made outstanding contributions to the fields of molecular biology and biotechnology. Our group recently developed a novel powerful fluorescent immunosensor strategy named Quenchbody (Q-body), which has been applied to the detection of a range of antigens in a rapid, simple, and sensitive manner. However, there were some Q-bodies whose fluorescence response was limited, especially for detecting protein antigens. With the aim of improving this issue, here we made twelve types of Q-bodies incorporated with different number and position of TAMRA fluorophore in the single chain Fv of HyHEL-10, an anti-hen egg lysozyme antibody, as a model. By measuring the fluorescence intensity and its antigen dependency, it was revealed that VL-VH type Q-bodies labeled at a non-CDR loop region of the VL shows the highest fluorescence response. This position locates close to the quenching Trp35 in VL, while it is far from Trp residues in the bound antigen. This result clearly suggests the importance of dye position to maximize the fluorescence quenching and antigen-dependent de-quenching. The discovery may open a way to make many other Q-bodies with superior response.
Keywords: Quenchbody; single chain antibody; fluorescence quenching; fluorescent biosensor; hen egg lysozyme; photoinduced electron transfer; in vitro translation Quenchbody; single chain antibody; fluorescence quenching; fluorescent biosensor; hen egg lysozyme; photoinduced electron transfer; in vitro translation
This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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MDPI and ACS Style

Jeong, H.-J.; Ueda, H. Strategy for Making a Superior Quenchbody to Proteins: Effect of the Fluorophore Position. Sensors 2014, 14, 13285-13297.

AMA Style

Jeong H-J, Ueda H. Strategy for Making a Superior Quenchbody to Proteins: Effect of the Fluorophore Position. Sensors. 2014; 14(7):13285-13297.

Chicago/Turabian Style

Jeong, Hee-Jin; Ueda, Hiroshi. 2014. "Strategy for Making a Superior Quenchbody to Proteins: Effect of the Fluorophore Position." Sensors 14, no. 7: 13285-13297.



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