Biosensors DOI: 10.1002/smll.200700453 Functional Quantum-Dot/Dendrimer Nanotubes for Sensitive Detection of DNA Hybridization** Chuan Liang Feng, Xin Hua Zhong, Martin Steinhart, Anne-Marie Caminade, Jean Pierre Majoral, and Wolfgang Knoll* The fun cti onalization of nan otu bes (NTs) is an eff ective strategy towards design of new hybrid materials combining customizedproperties and aniso tropy . [1–2] Such mater ials have attracted con sidera ble int ere st for app lic ati ons such as bio cat aly sts, bio sensor s, and as pla tfo rms for bio separa - tion. [3–5] For example, quantum dots (QDs) exhibiting narrow emi ssi on ban dwi dth , pho toc hemica l sta bil ity , and hig h quantum yie ld have bee n incorp ora ted int o the walls ofNTs. [6] However, the strategies for producing QD-modified NTs [7–8] report ed up to now suff er fro m low effic iency ofchemical functi ona liz ation and a lac k of con tro l over the spatial assembly of the QDs. Layer-by-layer (LBL) deposi- tion, [9] which involves the successive deposition of oppositely charg ed polye lectr olyte s, allows genera tion of functional multilayer systems with high precision, even onto complex subs tr at es such as nanopa rticles [10] and nanoporous matrices. [11–12] Nano parti cles bearin g charg ed ligan ds can easily be inco rporat ed into multilayer systems, [13] and the rational assemb ly of dif ferent-si zed QDs in LBLstruc tur es can yield so-called ‘‘nanorainbows’’ that emit white light. [14] By controlling the distance between the layers of different-sized QDs funnel-like bandgap profiles can be realized, [15–16] which can show rapid and efficient fluorescence resonance energy transfer (FRET) along the bandgap gradient. [17] A configuration that would be particularly advantageous for sensing consists of functionalized nanotubes aligned within the pores of a porous membrane, such as self -order ed nan opo rou s alumin a (anodi c alu minum oxide, AAO). [18] Wh ereas the synt hesi s of tubular nanost ructur es us ing such porous materials as shape- de fining molds is well- established, [19] the rational generation of complex functional- wall architectures has remained a challenge. Here, a strategy for the design and the fabrication of QD/dendrimer composite NTs inside AAO membranes with a pore diameter of 400 nm, a lattice constant of 500 nm, and a pore depth of 100 mm is reported. The arrays of aligned QD/dendrimer composite NTs enabled the dete ct ion of DNA hybr idization wi th signifi cantl y enhanced sensit ivity . The high speci fic surfa ce ar ea of the pore wall s of the AAO me mbranes (whi ch amounts to about 70 00 0 mm 2 pe r 10 mm 2 of membra ne sur fac e) combined with graded-bandgap architectures permits efficient energy transfer to the inner surfaces of the NTs, onto which si ngle-s tr ande d pr obe DNA is gr afte d. Emiss ion fr om dye -la bel ed tar get DNAcan thu s be pro bed wit h exc ept ion all y high sensitivity and selectivity after its hybridization to the probe DNA. Globular, fourth-generation N,N-disubstituted hydrazine phosphorus-contain ing dendrimers [20] hav ing 96 ter min al groups wit h either cationic [G 4 (NH þ Et 2 Cl À ) 96 ] (G 4 þ ) or anionic [G 4 (CH–COO À Na þ ) 96 ] (G 4 À ) end groups were used as the matrix forming the walls of the NTs. [21] As demon- str ate d pre viousl y, tub ula r nan ost ruc tur es can eas ily be obt ain ed by suc ces siv ely deposi tin g opp ositely cha rge d dendrimers into AAO templates. [22] The thickness of bilayers of linear polyelectrolytes deposited into nanoporous hosts is kno wn to sig nifican tly exceed tha t obt ained on smooth sub str ate s and is dif ficu lt to con trol. [11] The well- define d molec ular archi tectu re of dendr imeric polye lectr olyte s and their globula r shape allows us to overc ome these drawb acks, and hence, dendrimers are ideal components with which to build compartmentalized NT walls. [23] LBL assembly of QD/dendrimer multilayer systems was mon itored in situ by sur face plasmon resonance (SP R) spectroscopy, using planar Au model substrates coated with 3-mercaptopropionic acid (3-MPA). We consecutively depos- ited multilayers of G 4 þ dendrimers and QDs with an emission wavelength l ¼614 nm (QDs 614 ). The resulting kinetic mode SPR curves are shown in Figure 1a. The addition of each layer led to a regular increase of the reflectivity. The accumulated shift of the resonance angles was converted into the geometric film thickness by assuming a refractive index ofn %1.5 for the dendrimers [24] and ofn %2.7 for the QDs [25] (Figure 1b). The mean increase in the thickness of the multilayer system was determined to be Dd %2 nm for an additional dendrimer layer and Dd %5.5 nm for an additional QD 614 layer. The formation of multilayers containing a gradient assembly consisting ofQDs emitting at l ¼561 nm (QDs 561 ), QDs emitting at l ¼594 nm (QDs 594 ), and of QDs 614 on 3-MPA-coated Au substrates was also monitored by SPR. The accumulated shift of the resonance angles associated with the deposition of additional layers of the different QD species is shown in Figure 1c. The thickness of the multilayer system increased by Dd %2 nm for every additional dendrimer layer deposited, by Dd %6.5 nm for every additional QD 561 layer, by Dd %6.1 nm for every additional QD 594 layer, and by Dd %5.5 nm for every QD 614 communications [ Ã ] Prof. W. Knoll, Dr. C. L. Fe ng Max Planck Institute for Polymer Research Ackermannweg 10, 55128 Mainz (Germany) E-mail: knoll@mpip-main z.mpg.de Prof. X. H. Zhong Department of Chemistry East China University of Science & Technology 200237 Shanghai, (P.R. China) Dr. M. Steinhart Max Planck Institute of Microstructure Physics Weinberg 2, 06120 Halle (Germany) Prof. J. P. Majoral, Dr. A. M. Caminade Laboratoire de Chimie de Coordination Centre National de la Recherche Scientifique 205 Route de Narbonne, 31077 Toulouse Cedex 4 (France) [ ÃÃ ] Helpf ul discussions with Prof. T. Basche ´ , technical support by K. Sklarek, and funding by the German Research Foundation (SFB 625: ‘‘Von einzelnen Moleku¨len zu nanoskopisch strukturierten Materialien’’; SPP 1165 (STE 1127/6-3); CERC3 (Mu 334/22-2)) are gratefully acknowledged . : Supporting Information is available on the WWW under http:// www.small-journal.com or from the author. 566 ß 2008 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim small 2008, 4, No. 5, 566–571
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Chuan Liang Feng et al- Functional Quantum-Dot/Dendrimer Nanotubes for Sensitive Detection of DNA Hybridization
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8/3/2019 Chuan Liang Feng et al- Functional Quantum-Dot/Dendrimer Nanotubes for Sensitive Detection of DNA Hybridization
Dr. M. SteinhartMax Planck Institute of Microstructure Physics
Weinberg 2, 06120 Halle (Germany)
Prof. J. P. Majoral, Dr. A. M. Caminade
Laboratoire de Chimie de Coordination
Centre National de la Recherche Scientifique
205 Route de Narbonne, 31077 Toulouse Cedex 4 (France)
[ÃÃ] Helpful discussions with Prof. T. Basche, technical support by K.Sklarek, and funding by the German Research Foundation (SFB625: ‘‘Von einzelnen Molekulen zu nanoskopisch strukturiertenMaterialien’’; SPP 1165 (STE 1127/6-3); CERC3 (Mu 334/22-2)) aregratefully acknowledged.
: Supporting Information is available on the WWW under http://www.small-journal.com or from the author.
sensitivity resulting from enhanced Cy5 emission is observed.
The detection limit for DNA hybridization inside the NTs2
was studied by varying the concentration of Cy5-labeled targetDNA. Concentrations ranging from 100 nM to 100 f M were
applied and PL spectra were recorded with an excitation
wavelength of l¼ 460 nm (Figure 4d). A detection limit as low
as 100 f M (Figure 4d (5)) was found, and even for these low
concentrations the emission of the Cy5 could clearly be
separated from the background signal (Figure 4d (6)). To
evaluate the selectivity of the NTs2 functionalized with probe
DNA, a solution containing mismatch 2 (MM2) Cy5-labeled
target DNA was also studied. No hybridization or
nonspecific adsorption occurred, as evidenced by the absence
of Cy5 emission in the corresponding PL spectra (Supporting
Information, Figure 3). Therefore, NTs2 allow the detection of
DNA hybridization with both
significantly enhanced sensitivity
and high selectivity.
Taking into account the strong
dependence of the energy-transfer
efficiency on the distance between
donor and acceptor,[33] the chain
length of the probe DNA is a key
to successful sensing of the hybri-
dization with Cy5 labeled target
DNA. Figure 4e shows the PL
spectra after hybridization to
probe DNA of different chain
lengths. The PL intensity of the
Cy5 signal significantly decreases
as the distance between the chro-
mophores and the QD614 layer at
the inner surface of the NTs2
increases with an increased num-
ber of nucleotides. The strongest
PL intensity of Cy5 is observed forthe 30-mer probe DNA (Figure 4e
(4)). For the 50-mer probe DNA
with 20 additional thymines incor-
porated as spacers (between
the NH2 group grafted onto the
NTs2 and the 15-mer recognition
sequenceattheoppositeendofthe
probe), a much weaker PL inten-
sity of Cy5 is registered (Figure 4e
(3)). The lowest PL intensity is
detected with probe DNA having
80 nucleotides (with a total of 65
thymines spacers) in the strand(Figure 4e (2)).
For the detection of DNA
hybridization, NTs containing
cascaded energy transfer archi-
tectures have two major advan-
tages. Firstly, a much larger signal
amplification of the Cy5 emission
was observed as compared to
one-color systems based on single
QDs.[34] Secondly, although the
detection sensitivity of the NTs2 is not better than that
reported for other sensor systems, for example, nanoparticle-
based assays,
[35]
it should be possible to further optimize theperformance of QD/dendrimer NTs with graded bandgap
architectures by tuning the QD emission, by the incorporation
of additional QD species, and by adjusting the distance
between the QDs and organic dyes.
In conclusion, an efficient approach to the rational
assembly of different QD species in the walls of QD/
dendrimer composite NTs by LBL deposition has been
reported. Directed FRET through the graded bandgap
structure resulted in significantly enhanced detection of
DNA hybridization in the NTs combined with high selectivity.
Arrays of QD/dendrimer nanotubes aligned in AAO
membranes are particularly suitable for sensing because the
Figure 4. Detection of DNA hybridization. a) Schematic diagram displaying the direction of the energy
transfer from the QDs to the chromophore Cy5 attached to the target DNA after hybridization in the
NTs2. b) SPR kinetic scan during the immobilization of 30-mer probe DNA on smooth model
substrates. The inset shows the hybridization of Cy5-labeled complementary target DNA (15-mer) withprobe DNAinvestigated by SPFS. c) Normalized PL spectra of 1) NTs2 beforehybridization and 2) NTs2
after hybridization with Cy5-labeled target DNA using an excitation wavelength of l¼460 nm,
3) normalized PL spectra of the NTs2 before hybridization, and 4) after hybridization with Cy5-labeled
target DNA using an excitation wavelength of l¼ 630 nm. The same excitation intensity was used for
both excitation wavelengths. d) NormalizedPL spectra of the NTs2 after hybridization with Cy5-labeled
targetDNA usingsolutions with concentration of 1) 100 nM, 2 ) 1 0 nM, 3 ) 1 nM,4)100pM,and5)100f M.
e) Normalized PL spectra of the NTs2 1) before and 2-4) after hybridization of Cy5-labelled target DNA
to probe DNA with different strand lengths: 2) 80-mer; 3) 50-mer, and 4) 30-mer (excitation