Analytical Sciences

Abstract − Analytical Sciences, 32(3), 367 (2016).

Improvement of the Mutation-Discrimination Threshold for Rare Point Mutations by a Separation-Free Ligase Detection Reaction Assay Based on Fluorescence Resonance Energy Transfer
Kenta HAGIHARA, Kazuhiko TSUKAGOSHI, Chinami NAKAJIMA, Shinsuke ESAKI, and Masahiko HASHIMOTO
Department of Chemical Engineering and Materials Science, Faculty of Science and Engineering, Doshisha University, 1-3 Tataramiyakodani, Kyotanabe, Kyoto 610-0321, Japan
We previously developed a separation-free ligase detection reaction assay based on fluorescence resonance energy transfer from a donor quantum dot to an acceptor fluorescent dye. This assay could successfully detect one cancer mutation among 10 wild-type templates. In the current study, the mutation-discrimination threshold was improved by one order of magnitude by replacing the original acceptor dye (Alexa Fluor 647) with another fluorescent dye (Cyanine 5) that was spectrally similar but more fluorescent.