Fluorescence Reporting Based on FRET Between Conjugated Polyelectrolyte and Organic Dye for Biosensor Applications - Advanced Fluorescence Reporters in Chemistry and Biology - page 426

is bound to the resulting dsDNA, giving rise to high fluorescence. Addition of CCP
into the solution results in complex formation between CCP/dsDNA/intercalating
dye, subsequently leading to the amplified dye emission. In contrast, in the presence
of noncomplementary ssDNA strands, the fluorescence of intercalating dye is very
weak because of the inefficient FRET and the intrinsically low quantum yield of
free interlacing dye. However, this assay scheme is strongly dependent on the
choice of intercalating dye as well as CCP. For instance, neglectable FRET was
observed when ethidium bromide (EB) was used as the intercalating dye and 1I as
the donor [ 55 ], while sufficient FRET occurred between DNA-bound thiazole
orange (TO) and 1I [ 56 ]. It was suggested that the arrangement of donor and
acceptor transition dipole moments played an important role in controlling such
FRET processes [ 57 ]. In addition, our recent work shows that attachment of TO into
the side chain of 1I affords an efficient multicolor light-up probe (PFPTO) that can
distinguish dsDNA from ssDNA in serum media, which benefits from the stronger
affinity of TO toward dsDNA as compared to other biomolecules [ 58 ]. As shown in
Scheme 6 , in the absence of DNA, the fluorescent color of PFPTO solution is blue.
The blue fluorescence retains in the presence of ssDNA, while it gradually changes
to brown with increasing [dsDNA]. Consequently, naked-eye discrimination of
dsDNA from ssDNA in a mixed biological sample is feasible by using PFPTO as
a multicolor indicator. The dsDNA concentration can be qualitatively estimated
according to fluorescent color change of the polymer solution. This finding also
indicates that Scheme 5 could work in biological media.
On the basis of Scheme 5 , we recently realized label-free sequence-specific
DNA detection with SNP selectivity with the aid of S1 nuclease [ 59 ]. In this
assay, 1Br and TO are chosen as the energy donor and acceptor, respectively.

[dsDNA] / nM

0 1.2 2.4 3.6 4.8 6.0

Scheme 6 Photographs of fluorescence for PFPTO solutions at [RU] ¼ 2 m m in the presence of
dsDNA with (DNA) ranging from 0 to 6.0 nM at intervals of 1.2 nM in 1 PBS containing 10 vol

% serum under a hand-held UV lamp with l ¼ 365 nm

max