Squaraine-based polymer dots with narrow, bright near-infrared fluorescence for biological applications

Article abstract of DOI:10.1021/ja5123045

This article describes the design and development of squaraine-based semiconducting polymer dots (Pdots) that show large Stokes shifts and narrow-band emissions in the near-infrared (NIR) region. Fluorescent copolymers containing fluorene and squaraine units were synthesized and used as precursors for preparing the Pdots, where exciton diffusion and likely through-bond energy transfer led to highly bright and narrow-band NIR emissions. The resulting Pdots exhibit the emission full width at half-maximum of ～36 nm, which is ～2 times narrower than those of inorganic quantum dots in the same wavelength region (～66 nm for Qdot705). The squaraine-based Pdots show a high fluorescence quantum yield (QY) of 0.30 and a large Stokes shift of ～340 nm. Single-particle analysis indicates that the average per-particle brightness of the Pdots is ～6 times higher than that of Qdot705. We demonstrate bioconjugation of the squaraine Pdots and employ the Pdot bioconjugates in flow cytometry and cellular imaging applications. Our results suggest that the narrow bandwidth, high QY, and large Stokes shift are promising for multiplexed biological detections.

Full text of DOI:10.1021/ja5123045

Article
pubs.acs.org/JACS
Squaraine-Based Polymer Dots with Narrow, Bright Near-Infrared
Fluorescence for Biological Applications
I-Che Wu,†,‡ Jiangbo Yu,†,‡ Fangmao Ye,† Yu Rong,† Maria Elena Gallina,† Bryant S. Fujimoto,†
Yong Zhang,† Yang-Hsiang Chan,†,∥ Wei Sun,† Xing-Hua Zhou,† Changfeng Wu,§ and Daniel T. Chiu*,†
†Department of Chemistry, University of Washington, Seattle, Washington 98195, United States
§State Key Laboratory on Integrated Optoelectronics, College of Electronic Science and Engineering, Jilin University,
Changchun 130012, China
S
* Supporting Information
ABSTRACT: This article describes the design and development of squaraine-based
semiconducting polymer dots (Pdots) that show large Stokes shifts and narrow-band
emissions in the near-infrared (NIR) region. Fluorescent copolymers containing
fluorene and squaraine units were synthesized and used as precursors for preparing the
Pdots, where exciton diffusion and likely through-bond energy transfer led to highly
bright and narrow-band NIR emissions. The resulting Pdots exhibit the emission full
width at half-maximum of ∼36 nm, which is ∼2 times narrower than those of inorganic
quantum dots in the same wavelength region (∼66 nm for Qdot705). The squaraine-
based Pdots show a high fluorescence quantum yield (QY) of 0.30 and a large Stokes
shift of ∼340 nm. Single-particle analysis indicates that the average per-particle
brightness of the Pdots is ∼6 times higher than that of Qdot705. We demonstrate
bioconjugation of the squaraine Pdots and employ the Pdot bioconjugates in flow
cytometry and cellular imaging applications. Our results suggest that the narrow
bandwidth, high QY, and large Stokes shift are promising for multiplexed biological detections.
possess extraordinary brightness, fast emission rates, good
photostability, and minimal toxicity to biological cells and
tissues.16−22 There are recent efforts in developing Pdots with
NIR emission as fluorescent probes.23−25 One strategy is based
on encapsulation of a hydrophobic NIR dye in Pdots with visible
emission.23,24 Upon light excitation, the Pdots transfer the energy
efficiently to the dopant dye, resulting in quenching of visible
emission with concomitant NIR emission. However, this dye-
doping approach can have a dye leakage problem in applications
that occur over relatively long periods of time and during
storage.20,23,25 Another method is to blend a NIR-emitting
polymer as an acceptor with several semiconducting polymers
such as red, green, and blue-emitting polymers as donors that
work together via cascade fluorescence resonance energy transfer
(FRET) to generate NIR-emitting Pdots.25 The design and
synthesis of these polymers for cascade FRET are relatively
complicated, and much work must be done to optimize the
blending ratios of the semiconducting polymers to get high
fluorescence QY and efficient energy transfer.
INTRODUCTION
■
Near-infrared (NIR) fluorescent probes have been widely used
in biological and biomedical research because light in this
wavelength region has a large penetration depth in tissue and
causes minimal cellular autofluorescence.1 Organic dyes are the
most commonly used NIR fluorescent probes, despite their
non-optimal brightness and photostability. They are extensively
used in fluorescence imaging and assays because of their well-
established labeling protocols and their molecular dimen-
sions.2−8 However, the use of organic NIR probes for bioimaging
applications encounters several difficulties. For example, organic
NIR dyes usually consist of large, hydrophobic conjugated
systems that may lead to aggregation in physiological environ-
ments, which can dramatically reduce their emission quantum
yield (QY) and broaden their emission spectra as compared with
dyes emitting in the visible range.9 In addition, the Stokes shifts
for organic NIR dyes are generally very small, which is not
optimal for imaging applications because cross-talk between the
excitation light and fluorescence can make it difficult to capture
the pertinent signals.9 In the past decade, inorganic semi-
conductor quantum dots (Qdots) were developed as NIR probes
because they offer significantly higher photostability and
brightness compared to conventional dyes.10−12 The potential
leakage of heavy metal ions from the Qdots and the subsequent
cell toxicity, however, are critical concerns for biological
applications.12−15
Many biological applications require that the fluorescent
probes possess narrow-band emissions for multiplexed detec-
tions. For example, in a wavelength-and-intensity multiplexing
scheme for optical encoding, the number of codes increases
greatly with increasing number of color and intensity levels.
Previous studies suggest that it is better to use more colors, rather
Semiconducting polymer dots (Pdots) with small sizes
(<30 nm) are a new class of highly fluorescent probes that
Received: July 23, 2014
© XXXX American Chemical Society
A
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than more intensity levels, in order to increase the number of
usable codes.26 Therefore, development of narrow, bright NIR
Pdots remains an important task for widespread biological
applications. This article describes the design and synthesis of
squaraine-based Pdots that show large Stokes shifts and narrow-
band emissions in the NIR region. The squaraine Pdots exhibit
an emission bandwidth of ∼36 nm (full width at half-maximum,
fwhm), about 2 times narrower than those of Qdots in the same
wavelength region. To our knowledge, this is the narrowest
emission bandwidth among various Pdot species reported so far.
The narrow bandwidth, high QY, and large Stokes shifts of the
squaraine-based Pdots are promising for multiplexed biological
detections.
the Pdots. From the fluorescence spectra (Figure 1), all the
copolymers in THF solutions show two emission peaks, around
420 and 700 nm, respectively. The former originates from the
polyfluorene backbone and the latter is from the squaraine
derivatives. The intensity ratio (R) of the NIR emission peak to
the blue emission peak increases with increasing feeding ratios of
squaraine (Table S1). When these polymers were prepared into
Pdots, the nanoparticles in aqueous solution showed similar
trend in spectral evolution with dramatically increased NIR
emissions relative to the blue emission. However, the QY of the
NIR emission were also dramatically decreased with the higher
feeding ratio of squaraine monomers.
When the Pdots were excited with 405 nm light, we observed
strong NIR emission, despite the fact that there was almost no
spectral overlap (Figure 2) between the emission of the energy
donor (polyfluorene) and the absorption of the acceptor/emitter
(squaraine monomer). Energy transfer between molecules
without significant spectral overlap has been reported.27,28 We
believe the mechanism for the efficient energy transfer from
polyfluorene to squaraine likely is not based on FRET. Rather,
energy transfer likely occurred first through exciton diffusion
along the fluorene polymer backbone29−31 followed by through-
bond energy transfer27,28 to the squaraine monomer.
RESULTS AND DISCUSSION
■
Scheme 1 shows the chemical structures of the squaraine-based
polymers (PFS1 and PFS2) and the scheme for forming Pdot
Scheme 1. Chemical Structures of PFS1, PFS2, and PS-PEG-
a
COOH Polymers
Fluorescence lifetime measurements (Figure 2c,d) are helpful
for clarifying the mechanism of energy transfer processes.
We measured the fluorescence lifetimes of different donors and
acceptors in blended squaraine Sq1/PFO and PFS1 Pdots
(Figure 2c). From the fitted lifetime decay curves, the lifetime
values were 0.27 and 0.08 ns for fluorene in blended squaraine
Sq1/PFO Pdots and in PFS1 Pdots, respectively; for squaraine in
blended squaraine Sq1/PFO Pdots and in PFS1 Pdots, the values
were 2.0 and 2.24 ns, respectively. These values indicate fluo-
renes in PFS1 Pdots have a significantly shorter fluorescence
lifetime than those in the blended squaraine Sq1/PFO Pdots,
which is comparable to the published values for PFO Pdots. This
result suggests energy transfer from the fluorene donor to the
squaraine acceptor in PFS1 Pdots was much more efficient than
that in the blended Pdots; the enhanced intraparticle energy
transfer is the dominant factor that made PFS1 Pdots strong NIR
emitters. Similarly, in PFS2 Pdots, the luminescence lifetime of
fluorene donor was also shortened to 0.08 ns, which indicates
there was also an enhanced energy transfer in these Pdots. In
PFS2 Pdots, the NIR emission from the fused squaraine acceptor
also had a longer lifetime of 2.12 ns.
a
The schematic depicts the process of Pdot preparation, bioconjuga-
tion, and cellular labeling.
Table S1 summarizes the photophysical properties of the two
sets of Pdots with different monomer ratios. It is clear that the
fluorescence QY decreased with increased squaraine concen-
trations. For example, the QY of PFS1 Pdots decreased from
30% to 2% when the molar ratio of the squaraine chromophore
was increased from 1.5% to 10%. Similarly, the PFS2 Pdots
exhibited a decrease in QY from 17% to 1% as the molar con-
tent of benzene-fused squaraine increased from 1.5% to 10%.
We attribute this phenomenon to aggregation of the squaraine
chromophores in Pdots, consistent with the slight red-shift
observed in the Pdot fluorescence spectra. From these experi-
ments, we determined that Pdots consisting of 1.5% squaraine
offered the best overall performance in terms of the emission
bandwidth and brightness.
bioconjugates. The squaraine units were chosen because of their
strong absorption and narrow emission band centered in the NIR
region (700−720 nm). Two types of squaraine-based units were
synthesized and incorporated into the polyfluorene backbone
with different squaraine−fluorene ratios (squaraine monomer for
polymer PFS1 and benzene fused squaraine monomer for
polymer PFS2). The detailed synthesis procedure of the
squaraine monomer and polymer PFS1 is included in Schemes
S1 and S2 (Supporting Information). To further shift the
emission peak to the NIR, a benzene ring fused squaraine
monomer was also designed and prepared for polymer synthesis.
Benzene fused squaraine shows increased conjugation length
that generally produces a red-shifted emission. The synthesis
procedure for this fused squaraine monomer and polymer PFS2
is shown in Scheme S3 and S4 (Supporting Information).
Optimization of Monomer Ratio and Polymer Compo-
sition. We systematically studied the optical properties of the
Pdots as we varied the monomer ratios, as squaraine
concentration greatly affects the emission spectra and QY of
Colloidal and Photophysical Properties of Optimized
Pdots. Figure 3 shows the spectroscopic and colloidal properties
for the final optimized form of the two types of Pdots. An average
diameter of 18 nm was determined for both Pdots by dynamic
light scattering (DLS, Figure 3e,f), and further confirmed by
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Figure 1. (a) Emission spectrum for squaraine monomer (Sq1) in THF (λex = 600 nm, green), and emission spectra of PFS1 polymers in THF solution
with different molar ratios of acceptor squaraine (λex = 405 nm): 1.5% (black), 3% (red), 7% (blue), 9.4% (cyan), and 19% (magenta). The inset shows
the enlarged region of the fluorescence spectra of 1.5% (black) and 3% (red) PFS1 polymers in the range of 420−550 nm. (b) Emission spectrum for
fused squaraine monomer (Sq2) in THF (λex = 620 nm, green), and emission spectra of PFS2 polymers in THF solution with different molar ratios of
acceptor fused squaraine (λex = 405 nm): 1.5% (black), 5% (red), and 10% (blue). The inset shows the enlarged region of the fluorescence spectra of
1.5% (black) and 5% (red) PFS2 polymers in the range of 420−550 nm. (c) Emission spectra of different PFS1 Pdots made from the above PFS1
polymers in H2O (λex = 405 nm): 1.5% (black), 3% (red), 7% (green), 9.4% (blue), and 19% (magenta) of acceptor squaraine. (d) Emission spectra of
different PFS2 Pdots made from the above PFS2 polymers in H2O (λex = 405 nm): 1.5% (black), 5% (red), and 10% (blue) of fused squaraine acceptors.
transmission electron microscopy (TEM, Figure 3c,d). From
the absorption spectra, both Pdots showed strong absorption
peaks around 375 nm due to the polyfluorene backbone. The
fluorescence spectra show strong emission centers at 693 nm for
PFS1 Pdots, and 711 nm for PFS2 Pdots. Both Pdots show large
Stokes shifts of over 320 nm and exhibit narrow emission band-
widths (fwhm ∼36 nm), which is comparable to or narrower than
most NIR organic dyes, and about 2 time narrower than that of
Qdots emitting in similar wavelength region (∼66 nm for
commercially available Qdot705). Because the emission spectra
of Pdots show a tail at longer wavelengths, we also compared the
full width at 10% of maximal intensity between PFS1 Pdot
and Qdot705 (Figure S1); here, Pdot is still ∼37 nm narrower
than Qdot. The narrow emission bandwidth is important for
preventing emission cross-talk among probes, particularly for
multicolor labeling and multiplexed biological detections.
Single-Particle Brightness Comparison. For using Pdots
as fluorescent labels, the final brightness of Pdots can be
estimated by the product of the peak absorption cross-section
and the fluorescence QY of the NIR emission of the Pdots. The
absorption cross-section is the light-harvesting capability of a
fluorescent probe at a given wavelength, which can be
determined from the absorption spectra. An analysis of the
UV−vis spectra at known PFS1 and PFS2 Pdot concentrations
shows the per-particle absorption cross sections of the Pdots
(18 nm in diameter) at 405 nm were 2.4 × 10−13 cm2 for PFS1
Pdots and 1.9 × 10−13 cm2 for PFS2 Pdots, which are close to the
reported values of other types of Pdots.16,18,32 We performed
single-particle imaging to experimentally compare the single-
particle brightness between Qdot705 and the Pdots under
identical excitation and collection conditions. The PFS1 Pdots,
PFS2 Pdots, and Qdot705 were excited with a 405 nm laser, and a
long-pass filter of 685 nm was used. Figure 4 shows the single-
particle images of the three nanoparticles; intensity histograms
were obtained by analyzing hundreds of nanoparticles from each
sample. We found PFS1 Pdots were about six times brighter than
Qdot705 and PFS2 Pdots were about three times brighter than
Qdot705, consistent with their measured photophyiscal
parameters.
Cellular Labeling, Flow Cytometry, and Fluorescence
Imaging. To use the NIR-emitting Pdots for biological labeling
and targeting, we functionalized the Pdot surface with carboxyl
groups and carried out bioconjugation with streptavidin using
EDC-catalyzed coupling as described in our previous reports
(Scheme 1).18−20,25,32 Pdot-streptavidin (Pdot-SA) was used to
label cell-surface markers on MCF-7 breast cancer cells. The cells
were sequentially incubated with biotinylated primary anti-
EpCAM antibody and Pdot-SA probes. Flow cytometry was
performed to examine the specificity and brightness of labeling
by the Pdot-streptavidin probes (Figure 5). The labeled cells
exhibited stronger fluorescence compared with the control
samples, indicating the two Pdots effectively and specifically
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Figure 2. (a) Absorption spectra of squaraine monomer (Sq1) in THF (red dash) and PFS1 polymer in THF (violet dash), and emission spectra of PFO
polymer in THF (blue solid), PFS1 polymer in THF (red solid), and PFO polymer mixed with 1.5% (in molar ratio) Sq1 monomer in THF (cyan dash-
dot). (b) Absorption spectra of Sq1 monomer doped in amphiphilic PS-PEG-COOH (20 wt%) polymer nanoparticle in H2O (red dash), PFS1 Pdots in
H2O (violet dash-dot), and PFO polymer blended with 1.5% (in molar) Sq1 monomer Pdots in H2O (black dot), and emission spectra of PFO Pdots in
H2O (blue solid), PFS1 Pdots in H2O (burgundy solid), and blended Pdots prepared from PFO polymer mixed with 1.5% (in molar ratio) Sq1
monomer (red solid). (c) Instrument response function (IRF) curve (black), lifetime decay curves of the fluorescence from blended Sq1/PFO Pdots in
H2O, which include the lifetime decay for the fluorescence from fluorene groups of PFO polymer in Pdots (blue) and from squaraine monomer Sq1 in
Pdots (magenta), and lifetime decay curves of the fluorescence from PFS1 Pdots in H2O, which include the lifetime decay for the fluorescence from
fluorene groups in PFS1 Pdots (cyan) and from squaraine groups in PFS1 Pdots (red). (d) Lifetime decay curves of the fluorescence from fluorene
groups in PFS2 Pdots in H2O (blue) and from squaraine groups in PFS2 Pdots in H2O (red). All emission spectra were obtained at an excitation of λex =
380 nm. All fluorescence lifetime measurements were obtained at an excitation from a laser tuned to λex = 375 nm.
Table 1
b
c
d
size
(nm)
λem
(nm)
σ
(×10−13
QY
fwhm
(nm)
τ
sample
cm2)
(%)
(ns)
Qdot705
PFS1
13
705
693
711
0.32
2.4
41
30
17
66
36
36
6533
2.24
2.12
a
18
a
PFS2
18
1.9
a
b
Average size measured by DLS. Single-particle absorption cross-
c
section at 405 nm. Quantum yield was measured at a 405 nm
excitation wavelength. Fluorescence lifetime.
d
labeled EpCAM receptors on the cell surface with excellent
brightness. The negative control samples were prepared under
identical conditions as the positive ones but without primary
biotinylated antibodies. We compared the PFS1 and PFS2 Pdots-
SA with commercial Qdot705-SA under the same labeling
concentrations and identical experimental conditions (Figure 5).
The results indicate PFS1-SA and PFS2-SA Pdots were about six
and three times brighter than that of Qdot705-SA, respectively,
which are consistent with the single-particle brightness measure-
ments shown in Figure 4. From the negative control samples, it
can be seen that both Pdot probes also exhibited less non-specific
binding to cells than the Qdot probe.
We further confirmed specific cellular labeling with PFS1
Pdot-SA and PFS2 Pdot-SA probes by confocal fluorescence
imaging (Figure 6). These results also show PFS1 (Figure 6a)
and PFS2 Pdot-SA (Figure 6b) probes effectively and specifically
labeled EpCAM receptors on the MCF-7 cell surface; in the
equivalent negative control experiments (data not shown) where
the primary biotinylated antibody was absent, no fluorescence
Figure 3. Absorption and emission spectra (λex = 380 nm) of (a) PFS1
and (b) PFS2 Pdots in water. TEM images for (c) PFS1 Pdots and (d)
PFS2 Pdots; scale bar is 50 nm. Histograms of size distribution measured
by DLS for (e) PFS1 Pdots and (f) PFS2 Pdots; both show an average
diameter of 18 nm.
was detected, again indicating little or no non-specific binding of
the Pdots to the cells.
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Figure 4. Single-particle fluorescence images of (a) Qdot705, (b) PFS1 Pdots, and (c) PFS2 Pdots obtained with a 405 nm laser using the same
excitation power and under identical detection conditions. Histograms of the distributions of single-particle brightness of (d) Qdots705, (e) PFS1 Pdots,
and (f) PFS2 Pdots. The black curves were obtained by fitting a log-normal function to the histograms, which gave mean brightness of 4500, 25 300, and
13 300 CCD counts for Qdot705 and PFS1 and PFS2 Pdots, respectively.
Figure 5. Flow cytometry measurements of the labeling intensity of MCF-
7 breast-cancer cells labeled with Qdot705-streptavidin, PFS1 Pdot-
streptavidin, and PFS2 Pdot-streptavidin in the presence (positive labeling,
P) and absence (negative control, N) of biotinylated primary antibody
against the cell-surface protein EpCAM. (a) Comparison between
Qdot705 and PFS1 Pdots; fluorescence of labeled cells was collected
with a 695/40 nm band-pass filter; excitation at 405 nm. (b) Comparison
between Qdot705 and PFS2 Pdots; fluorescence of labeled cells was
collected with a 710/20 nm band-pass filter; excitation at 405 nm.
Figure 6. Confocal fluorescence microscopy images of MCF-7 cells
labeled with (a) PFS1 Pdot-streptavidin via binding to biotinylated anti-
EpCAM primary antibody and (b) PFS2 Pdot-streptavidin via binding
to biotinylated anti-EpCAM primary antibody. From left to right:
fluorescence images, Nomarski images, and merged fluorescence and
Nomarski images; scale bars are 50 μm.
CONCLUSION
■
that of Qdot705. These superior performance features should
make PFS Pdots a valuable new addition in the near-IR region for
multiplex biological imaging and analysis.
In summary, we successfully developed two types of near-
infrared-emitting polymer dots with large Stokes shift based on
narrow-band emissive squaraine derivatives. Rather than FRET,
we believe that efficient exciton diffusion along the fluorene
polymer followed by through-bond energy transfer occurred in
the PFS Pdots. This mechanism enabled us to develop NIR-
emitting Pdots with large Stokes shift, high quantum yield, and
narrow emission bandwidth, even in the absence of significant
spectral overlap between energy donor and acceptor. Single-
particle brightness measurements and flow-cytometry experi-
ments both indicate that the PFS Pdots can be up to 6 times
brighter when compared with Qdot705, while the emission
bandwidths of the PFS Pdots are about 2 times narrower than
ASSOCIATED CONTENT
* Supporting Information
Materials, experimental details, and supplementary figures and
synthetic schemes. This material is available free of charge via the
Internet at http://pubs.acs.org.
■
S
AUTHOR INFORMATION
Corresponding Author
chiu@chem.washington.edu
■
E
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Journal of the American Chemical Society
Article
(25) Zhang, X.; Yu, J.; Rong, Y.; Ye, F.; Chiu, D. T.; Uvdal, K. Chem. Sci.
2013, 4, 2143.
(26) Han, M.; Gao, X.; Su, J. Z.; Nie, S. Nat. Biotechnol. 2001, 19.
(27) Lin, W.; Yuan, L.; Cao, Z.; Feng, Y.; Song, J. Angew. Chem., Int. Ed.
2010, 49, 375.
Present Address
∥Department of Chemistry, National Sun Yat-sen University, 70
Lien Hai Road, Kaohsiung, Taiwan 80424.
Author Contributions
‡I-C.W. and J.Y. contributed equally.
(28) Jiao, G.-S.; Thoresen, L. H.; Burgess, K. J. Am. Chem. Soc. 2003,
125, 14668.
Notes
(29) Lyons, B. P.; Monkman, A. P. Phys. Rev. B 2005, 71, 235201.
(30) Bruno, A.; Reynolds, L. X.; Dyer-Smith, C.; Nelson, J.; Haque, S.
A. J. Phys. Chem. C 2013, 117, 19832.
The authors declare the following competing financial
interest(s): J.Y., C.W., X.-H.Z., and D.T.C. have financial interest
in Lamprogen, which has licensed the described technology from
the University of Washington.
(31) Peckus, D.; Devizis, A.; Hertel, D.; Meerholz, K.; Gulbinas, V.
̌
Chem. Phys. 2012, 404, 42.
(32) Wu, C.; Hansen, S. J.; Hou, Q.; Yu, J.; Zeigler, M.; Jin, Y.;
Burnham, D. R.; McNeill, J. D.; Olson, J. M.; Chiu, D. T. Angew. Chem.,
Int. Ed. 2011, 50, 3430.
ACKNOWLEDGMENTS
■
We gratefully acknowledge support for this research from the
National Institutes of Health (R21CA186798) and the
University of Washington. C.W. acknowledges financial support
from “Thousand Young Talents Program” and the National
Science Foundation of China (Grant Nos. 61222508 and
61335001).
(33) Bharadwaj, P.; Novotny, L. Nano Lett. 2011, 11, 2137.
REFERENCES
■
(1) Tung, C. H.; Lin, Y. H.; Moon, W. K.; Weissleder, R. ChemBioChem
2002, 3, 784.
(2) Escobedo, J. O.; Rusin, O.; Lim, S.; Strongin, R. M. Curr. Opin.
Chem. Biol. 2010, 14, 64.
(3) Kovar, J. L.; Simpson, M. A.; Schutz-Geschwender, A.; Olive, D. M.
Anal. Biochem. 2007, 367, 1.
(4) Achilefu, S. Angew. Chem., Int. Ed. 2010, 49, 9816.
(5) Nesterova, I. V.; Verdree, V. T.; Pakhomov, S.; Strickler, K. L.;
Allen, M. W.; Hammer, R. P.; Soper, S. A. Bioconjugate Chem. 2007, 18,
2159.
(6) Kukrer, B.; Akkaya, E. U. Tetrahedron Lett. 1999, 40, 9125.
(7) Dilek, G.; Akkaya, E. U. Tetrahedron Lett. 2000, 41, 3721.
(8) Oswald, B.; Lehmann, F.; Simon, L.; Terpetschnig, E.; Wolfbeis, O.
S. Anal. Biochem. 2000, 280, 272.
(9) Brasseur, N.; Nguyen, T.-L.; Langlois, R.; Ouellet, R.; Marengo, S.;
Houde, D.; van Lier, J. E. J. Med. Chem. 1994, 37, 415.
(10) Chan, W. C. W.; Maxwell, D. J.; Gao, X.; Bailey, R. E.; Han, M.;
Nie, S. Curr. Opin. Biotechnol. 2002, 13, 40.
(11) Zheng, J.; Zhang, C.; Dickson, R. M. Phys. Rev. Lett. 2004, 93,
No. 077402.
(12) Michalet, X.; Pinaud, F. F.; Bentolila, L. A.; Tsay, J. M.; Doose, S.;
Li, J. J.; Sundaresan, G.; Wu, A. M.; Gambhir, S. S.; Weiss, S. Science
2005, 307, 538.
(13) Medintz, I. L.; Uyeda, H. T.; Goldman, E. R.; Mattoussi, H. Nat.
Mater. 2005, 4, 435.
(14) Somers, R. C.; Bawendi, M. G.; Nocera, D. G. Chem. Soc. Rev.
2007, 36, 579.
(15) Soo Choi, H.; Liu, W.; Misra, P.; Tanaka, E.; Zimmer, J. P.; Itty
Ipe, B.; Bawendi, M. G.; Frangioni, J. V. Nat. Biotechnol. 2007, 25, 6.
(16) Wu, C.; Bull, B.; Szymanski, C.; Christensen, K.; McNeill, J. ACS
Nano 2008, 2, 2415.
(17) Yu, J.; Wu, C.; Sahu, S. P.; Fernando, L. P.; Szymanski, C.;
McNeill, J. J. Am. Chem. Soc. 2009, 131, 18410.
(18) Wu, C.; Schneider, T.; Zeigler, M.; Yu, J.; Schiro, P. G.; Burnham,
D. R.; McNeill, J. D.; Chiu, D. T. J. Am. Chem. Soc. 2010, 132, 15410.
(19) Ye, F. M.; Wu, C. F.; Jin, Y. H.; Chan, Y. H.; Zhang, X. J.; Chiu, D.
T. J. Am. Chem. Soc. 2011, 133, 8146.
(20) Yu, J.; Wu, C.; Zhang, X.; Ye, F.; Gallina, M. E.; Rong, Y.; Wu, I.
C.; Sun, W.; Chan, Y.-H.; Chiu, D. T. Adv. Mater. 2012, 24, 3498.
(21) Rong, Y.; Wu, C.; Yu, J.; Zhang, X.; Ye, F.; Zeigler, M.; Gallina, M.
E.; Wu, I. C.; Zhang, Y.; Chan, Y.-H.; Sun, W.; Uvdal, K.; Chiu, D. T.
ACS Nano 2013, 7, 376.
(22) Wu, C. F.; Chiu, D. T. Angew. Chem., Int. Ed. 2013, 52, 3086.
(23) Jin, Y. H.; Ye, F. M.; Zeigler, M.; Wu, C. F.; Chiu, D. T. ACS Nano
2011, 5, 1468.
(24) Xiong, L. Q.; Shuhendler, A. J.; Rao, J. H. Nat. Commun. 2012, 3.
F
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