Biomedical Engineering Reference
In-Depth Information
based on the hybridization of target RNA with a capture probe attached to a solid
support (shown in Fig.
3.9
) [
34
]. 3′-termini biotinylated miRNA targets hybridized
with the corresponding complementary DNA probes, which were immobilized on
glass slides. Streptavidin-modified QDs were conjugated with the biotin-miRNA
to read out signal (Fig.
3.9
a and b). Analysis of a model system indicated that the
detection limit for analyzing miRNA was about 0.4 fmol and the detection dynamic
range spanned across two orders of magnitude, from 156 to 20,000 pM (Fig.
3.9
c).
Moreover, the method was applied to develop an assay for profiling 11 miRNA tar-
gets from rice (Fig.
3.9
d). However, the lack of orthogonal conjugation methods for
attaching miRNAs to QDs did not allow the use of different QD colors.
To achieve signal enhancement and multiplexed analysis, QD nanobarcode-
based microbead random array platform for accurate and reproducible gene expres-
sion profiling in a high-throughput and multiplexed format was developed as shown
in Fig.
3.10
[
35
]. Four different sizes (and thus four different fluorescent colors)
of Qdots, with emissions at 525, 545, 565, and 585 nm, are mixed with a polymer
Fig. 3.9
Analysis of miRNA by means of fluorescent QDs.
a
Principle of labeling miRNA at 3′
termini with biotin.
b
Organization of the streptavidin-labeled QDs on a DNA/miRNA duplex
bound to a glass support.
c
Fluorescence intensities detected upon analyzing different concentra-
tions of a target miRNA (
upper panel
) and derived calibration curve (
lower panel
).
d
Analysis
in an array format of 11 target miRNAs from rice. Reproduced with permission from Ref. [
34
].
Copyright 2005, Oxford University Press
