Biomedical Engineering Reference
In-Depth Information
◄
Fig. 3.16
Scheme for polymer coating of QD-TOPO (
a
) and the conjugation reaction with
amine-terminated oligonucleotides using EDC-coupling reagent (
b
). Specificity of QD-DNA
probes hybridization in situ to detect mRNA (Rp49) in fixed Drosophila S2 cells.
c
Control
(QD555 alone, no DNA attached).
d
QD555-DNA conjugate.
e
Control experiment with
QD555-noncomplementary DNA conjugate.
f
RNase A treatment 10 mg mL prior to cell fixa-
tion significantly decreased the FISH signal.
g
Representative single-cell FISH image separately
displaying a DAPI-stained cell nucleus and the QD-DNA conjugates (for Rp49) exclusively
located in the cytoplasm, and the merged image of the DAPI signal and QD555 signal.
Reproducibility was obtained from three separate experiments. The scale bar indicates 20 mm
(
c
-
f
) and 5 mm (
g
). Reproduced with permission from Ref. [
62
]. Copyright 2009, Wiley
of QDs in such DNA microarray is ascribed to the narrow multicolor emissions
under a single-source excitation. Meissner et al. developed a QD-embedded
microsphere-based fluid DNA microarray. CdSe/ZnS QDs embedded in polysty-
rene microspheres were labeled with DNA oligonucleotides for target capture,
and the result was recorded by a high-speed readout flow cytometer [
68
]. QD-
based cDNA microarray was also developed for single-nucleotide polymorphism
(SNP) mutation detection in the human p53 tumor suppressor gene and multial-
lele detections. The authors established a model that used a SNP located at amino
acid residue 248, on exon 7 of p53, which is one of the most common mutation
hot spots in p53 [
69
]. As shown in Fig.
3.17
, multicolor targeting of SNP muta-
tions in human oncogene p53 and of human hepatitis B and C viruses with differ-
ent DNA-QDs were realized, which displayed the great potential of DNA-QD
conjugates as efficient probes in cDNA microarrays for a ultrafast detection of a
great number of viral or bacterial pathogens simultaneously. However, this model
system was not extended to real samples such as blood [
70
].
Fig. 3.17
Multicolor targeting of SNP mutations in human oncogene p53 (rows
1
-
6
) and of
human hepatitis B and C viruses (rows
7
-
10
) with different DNA-QD probes. In panel
a
,
yellow
Dp53 g-QDs are targeted only toward mutations in human oncogene p53, but SNP detection is
not achieved. In panel
b
,
red
B3-QDs are targeted toward the hepatitis B virus sequence. In panel
c
,
yellow
C5-QDs are targeted only toward the hepatitis C virus sequence. Panel
d
is an overlay
of panels
a
-
c
. In both
red
and
yellow
channels, signal-to-noise ratio is >100, and no cross talk
is observed. Reproduced with permission from Ref. [
70
]. Copyright 2003, American Chemical
Society
