Article pubs.acs.org/JPCB
Modulated Fluorescence of Colloidal Quantum Dots Embedded in a Porous Alumina Membrane Hao Xu,† Li Li,† Otto Manneberg,† Aman Russom,† Kristinn B. Gylfason,‡ Hjalmar Brismar,† and Ying Fu*,† †
Science for Life Laboratory, Department of Applied Physics, Royal Institute of Technology, SE-106 91 Stockholm, Sweden Micro and Nanosystems, Royal Institute of Technology, SE-100 44, Stockholm, Sweden
‡
ABSTRACT: The fluorescence spectrum of CdSe core-CdS/ZnS shell colloidal quantum dots (QDs) embedded in porous alumina membrane was studied. Small peaks, superimposed on the principal QD fluorescence spectrum, were observed. Finite-difference time-domain simulation indicates that the QD point radiation emitting from within the membrane is strongly modulated by the photonic band structure introduced by the membrane pores, leading to the observed fine spectral features. Moreover, the principal QD fluorescence peak red-shifted when the optical excitation power was increased, which is attributed to QD material heating due to emitted phonons when the photoexcited electron and hole relax nonradiatively from high-energy states to the ground exciton state before fluorescence.
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from Whatman Ltd.,21 as a first step toward the QD-membrane technique. In contrast to nanoporous membranes, the Whatman membranes show basically no optical response (see below). However, porous materials show extraordinary light reflection and transmission due to their photonic band structure.22 We therefore expect modulation of the QD fluorescence by the photonic band structure of the Whatman porous membrane, when QDs are embedded in the membrane, since the wavelength of the QD fluorescence is comparable with the pore size of the membrane. Nanoporous membranes, on the other hand, are not expected to affect the QD fluorescence in the visible range because of the nanopore sizes.
INTRODUCTION Colloidal quantum dots (QDs) have been attracting more and more researchers in the biomedical imaging field in the past decade. 1−4 Compared to traditional organic dyes and fluorescent proteins, QDs of nano size (2−10 nm) show great application potential due to their unique optical properties,5−7 such as high photoluminescence quantum yields, narrow emission spectra, tunable energy bandgaps, simultaneous excitation of multiple fluorescent colors, and superior photostability (much less photobleaching). Moreover, QDs, especially water-dispersible QDs, have been successfully and stably conjugated to specific biological molecules,8−10 and further used for in vitro cell labeling and in vivo imaging purposes.11,12 On the other hand, porous alumina membranes, with high pore density, tunable pore size and distribution, low protein binding, and minimal autofluorescence, are extensively used as biomolecular filters13 and for separating particles in liquids,14 for, e.g., proteomics and disease diagnostics.15−18 A potential application could be combining QDs and the membrane separation technique into one composite system, where selectively QD-conjugated biomolecules are captured in a porous membrane, through the attachment of specific surface ligands around QDs. This would enable rapid identification and in vitro diagnosis, through the simple monitoring of the QD fluorescence signal. Fluorescence spectra of QDs, e.g., CdSe/ZnS QDs19 and Si QDs,20 have been studied and reported after embedding QDs into nanoporous anodic alumina membranes, showing an unmodulated QD fluorescence spectrum and a modulated fluorescence from the nanoporous alumina membranes. Nanoporous (pore diameter of 50 nm) alumina membranes have a broad emission peak ranging from 350 to 600 nm. In this work, we study the fluorescence of QDs embedded in porous aluminum oxide membranes (Anodisc 47, 6809−5522) with pore size 0.2 μm and average thickness 60 μm purchased © 2013 American Chemical Society
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EXPERIMENTAL METHODS
Sample Preparation. Water-dispersible CdSe-CdS/ZnS coremultishell QDs with an emission-peak wavelength of 607 nm at room temperature were made in-house by common chemical synthesis (see our earlier work3,23). More specifically, a mixture of CdO (13 mg, 0.1 mmol), trioctylphosphine oxide (TOPO) (0.65 g, 1.7 mmol), ntetradecylphosphonic acid (TDPA) (56 mg, 0.2 mmol), and octadecylamine (ODA) (0.35 g, 1.3 mmol) was put into a 50 mL three-neck flask which was vacuumed for 30 min at 100 °C. After that, the flask was heated to 330 °C under nitrogen flow to get a clear solution. The temperature was then lowered down to 260 °C and 12 mg Se dissolved in 0.4 mL tributylphosphine (TOP) was injected quickly into the reaction mixture. We kept the temperature at 260 °C for the QDs core growth, then switched off the heat supply for the reaction flask when the desired QD size was reached. When the temperature came down to 100 °C, 15 mL acetone was added, leading to a precipitation of the QDs. Centrifugation and decantation were carried out subsequently several times to remove residual precursors and purify the QD cores. The following procedure was performed to add shell structures to the CdSe core: 1 g ODA, 4 mL 1-octadecene Received: September 12, 2013 Revised: October 16, 2013 Published: October 17, 2013 14151
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(ODE) were put into the 50 mL three-neck flask which was vacuumed for 30 min at 100 °C. The CdSe cores synthesized before were dissolved in chloroform and quickly injected into the reaction solution, which was then heated to and kept at 230 °C under nitrogen flow; Cd−, S−, and Zn− precursors were injected into the reaction mixture sequentially to get the desired core/shell CdSe QDs. Finally, 3mercaptopropionic acid (3-MPA) was used to replace ODA on the oildispersible QDs to make them water dispersible. The following samples were prepared and studied: Sample A: untreated membrane. The membrane was immersed in immersion oil (Immersol 518F, Zeiss) on a microscope slide and covered by a coverslip. Sample B: QDs in dispersion. QDs dispersed in an aqueous continuous phase at a concentration of 100 nM were dropped into a circular area formed by nail polish on a microscope slide, and covered by a coverslip. Excitation light was focused on the center of the circle. Sample C: membrane with embedded QDs. The membrane was immersed in a 3 nM QD dispersion for 100 min to adsorb QDs. It was then put into water for 20 min, to wash away loosely attached QDs, and subsequently air-dried and immersed into oil on a microscope slide. Sample D: highly diluted oil-soluble CdSe core/shell QDs (at a concentration of 10 pM) distributed on a microscope slide by dip coating, then dried at ambient temperature. The porous structure of a representative membrane was characterized by scanning electron microscopy (SEM), at an acceleration voltage of 15 kV, using an FEI Nova NanoSEM 650 system. Figure 1a shows a typical SEM image of the membrane surface and Figure 1b a typical cross-section, confirming membrane specifications.
Figure 2. (a) Optical reflectance spectra from the microscope slide (dashed line), and the sample A membrane on a slide (solid line), under 488 nm laser excitation. (b) Fluorescence spectra of the sample B QD dispersion (dashed line, peak position 607 nm), and the sample C membrane with embedded QDs (solid line, peak position 610 nm), from the circular area of diameter of 1.4 μm indicated in (c), with different normalization factors for direct comparison.
membrane with embedded QDs (sample C, circular collection area with a diameter of 1.4 μm, as indicated in Figure 2c), with different normalization factors for the convenience of comparison, showing that when the fluorescence spectrum of the embedded QDs is collected from a large area, it is very similar to that of QDs in dispersion. However, a small redshift was observed for the embedded QDs, which will be discussed below. Figure 3a shows the fluorescence spectra obtained from a series of circular areas of diameter 0.21 μm on sample C (see inset). Small peaks, superimposed on the principal QD fluorescence spectra, were observed. Figure 3b demonstrates how the small peaks (fine spectral features) are averaged out when the observation area is increased gradually (diameter increasing from 0.14 to 1.4 μm). To understand the fine spectral features in Figure 3a and b, we dispersed highly diluted oil-soluble CdSe core/shell QDs (10 pM) on a microscope slide (sample D). The QDs were so diluted that blinking was observed under continuous wave excitation (for experimental details, see ref 3). As shown in Figure 4a, we chose four single QDs marked by small rings in Figure 4c and measured their florescence spectra from circular areas with the same diameter of 0.21 μm, i.e., the same circular size as in Figure 3a. We further obtained another four fluorescence spectra (see Figure 4b) from neighboring QDs enclosed in large ellipses in Figure 4c. It is easily observed that the measured fluorescence spectra of various QDs on the structure-less microscope slide surface (Sample D) are rather smooth, much alike the QDs in dispersion. Moreover, the observed dispersion of the peak positions of single QD spectra in Figure 4a is only around 10 nm, and the fluorescence peak in Figure 4b from two neighboring QDs splits into two peaks with a distance of approximately 6 nm, while the fine spectral features in Figure 3 extend over a spectral width of at least 70 nm. It is therefore concluded that the fine spectral features presented in Figure 3 should be attributed to the porous structure of the membrane. Finite-Difference Time-Domain Simulation. We employed the finite-difference time-domain (FDTD) method to study the fine spectral features in the fluorescence from QDs embedded in porous membranes. The porous structure of the
Figure 1. Typical SEM images of the Whatman aluminum oxide membrane. (a) The surface morphology shows that the diameter of the pores is about 0.2 μm. (b) A cross-section shows that the thickness of the membrane is about 55 μm. Characterization Method. All samples listed above were characterized using an LSM 780 confocal microscope (Carl Zeiss, Jena, Germany) with a Plan Apochromat 63 × 1.4 Oil DIC M27 objective, and a 32-channel GaAsP spectral detector, at an excitation wavelength of 488 nm. The optical power of the excitation laser was varied. The fluorescence emission from the samples was recorded over a bandwidth of 546−671 nm with a spectral step of 2.9 nm. Each imaging frame contains 128 × 128 pixels, and the pixel size is 70 × 70 nm2; thus, the imaging size is 9.0 × 9.0 μm2. The software ImageJ24 was used to read the fluorescence signals from the images acquired by the microscope. The mean value of the pixels in the selected area was used for analysis.
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RESULTS AND DISCUSSION Fine Spectral Features in the Fluorescence Spectra. We first measured the optical response of the microscope slide and the membrane. Figure 2a shows the reflectance spectra under 488 nm illumination from a clean microscope slide and a clean membrane on a slide (sample A), indicating negligible optical response from the membrane. Figure 2b presents fluorescence spectra of the QD dispersion (sample B) and a 14152
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Figure 5. (a) Geometry of the FDTD simulation. The annotated x, y, and r indicate the positions and radii of the pores. (b) Optical spectra of the light recorded by the planar detector. Red dashed line: the spectrum of the point source in immersion oil; solid lines: spectra with immersed porous-membrane structure, when the point source is situated at different positions (shown by value of d corresponding to the left structure).
from z = −0.6 to z = 0.1 μm, i.e., from inside the membrane to outside the membrane. The simulation space along the z direction was between z = −1 and z = 3 μm, terminated by perfectly matched layers (PMLs). A planar detector was placed at z = 2.2 μm, to record the electric field which consists of the direct light from the point source as well as the reflected light from the membrane, the same geometric configuration as that of sample C. The mesh size of the FDTD simulations was 6 nm. The simulated optical spectra are presented in Figure 5b. The detected light intensity from a point source located at d = 0.1 μm was almost twice that of a source located inside the membrane, i.e., at d ∈ (0.0, −0.6) μm. The higher intensity is due to the reflection from the membrane. Weak ripples were seen in the spectrum of d = 0.1 μm, and also appeared in the point source because of the finite wavelength range involved in the numerical FDTD calculations. Figure 5 shows that the light from the point source was strongly modulated when the source was positioned on the membrane surface and inside the membrane, i.e., at d ∈ (−0.6, 0.0) μm, resulting in fine spectral features similar to those observed experimentally in Figure 3. Similar fine spectral features, but at different wavelengths, were obtained in simulated spectra, when the positions and sizes of the four pores in the membrane were varied. This explains the disappearance of the fine spectral features in Figure 3b when the observation area was enlarged. The photonic band structure modulation is only observable when the QD is positioned on the membrane surface or inside a pore of the membrane. The effects are clearly observed in the spatial distributions in Figure 6 of the electric fields from QDs outside and inside the pore, respectively, at two different times (time step 200 and 400, corresponding to real times of 220 and 440 ps, respectively). In the FDTD computation domain, the xy plane is periodic so that in Figure 6a we observe not only the electric field from the QD at x = 0.15 μm but also the electric field from the QD at x = 0.75 μm at t = 200, while in Figure 6b and d we observe the interference between the electric fields from the two QDs. The refractive index of the membrane is relatively low, and thus the degree of the spatial variation in the refractive index is limited along the z direction, so that the light from the QD
Figure 3. Fluorescence spectra features of QDs embedded in a membrane (sample C). (a) Fluorescence spectra obtained from a series of circular areas of diameter 0.21 μm along the vertical direction (arrow). (b) Fluorescence spectra from concentric circular areas with diameters increasing from 0.14 to 1.4 μm.
Figure 4. (a) Fluorescence spectra of the single oil-soluble CdSe core/ shell QDs indicated by small rings in (c). (b) Fluorescence spectra of neighboring QDs enclosed by large ellipses in (c). (c) One image frame showing the spatial locations of the QDs whose fluorescence spectra are presented in (a) and (b).
membrane was optically represented by the spatial variation of the refractive index, i.e., n = 1.5 in porous areas filled with immersion oil and n = 1.7 in the alumina.25 Figure 5a shows the geometric structure of our FDTD simulations. The membrane structure was simulated by a film of 0.6 × 0.6 μm2 periodically extended in the xy plane, with a thickness of 0.6 μm (z ∈ (−0.6, 0.0) μm). This membrane patch was irregularly perforated with four pores of diameters around 0.2 μm. The QD fluorescence was simulated by a point Gaussian-pulse source with a wavelength range from 540 to 690 nm centered at 607 nm, as illustrated by the red dashed line in Figure 5b. The point source was positioned at (0.15, 0.15, d) μm, where d was varied 14153
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Figure 7. Fluorescence spectra vs excitation powers (from power 20% to 100% at a step of 10%) from sample C and QD dispersion (1/ 10000, 1/3600, and 1/4000 are the intensity scaling factors applied). Fluorescence spectra under excitations of different laser powers (a) from a large area enclosing 650 pixels (3.18 μm2); (b) from a small area enclosing 140 pixels (0.69 μm2). (c) Peak wavelength vs excitation power for large (solid stars) and small (hollow stars) areas. (d) Spectra from QD dispersion (sample B) under different power excitations 20%, 50%, and 100%.
Figure 6. Electric field distributions at time step 200 (a,c) and 400 (b,d). (a,b) The QD (marked as “+”) is positioned at d = 0.2 μm; (c,d) the QD (marked as “+”) is positioned at d = −0.3 μm. Dashed lines mark the boundaries of the simulated membrane and the edges of the pore in the membrane.
outside the pore is largely reflected (see Figure 6a,b). The short-range ordered pores in the xy plane modify the light propagation in that plane and induce photonic resonance modes,26,27 which are clearly observed in Figure 6b,d. Note that the edges of the point light source look cubic in Figure 6a which is caused by the cubic FDTD lattice. Moreover, the membrane thickness in the FDTD simulation is only 0.6 μm, while it is about 55 μm in reality. As seen in Figure 6, the photonic resonance modes appearing in the electric field distribution are largely caused by the refractive index modulation in the xy plane, and the light reflection from the membrane boundary at z = −0.6 μm is very limited. A thin membrane is therefore necessary to be simulated. On the other hand, FDTD can, in theory, handle the 55-μm-thick membrane by extending the computation time. However, a long-time calculation can easily result in numerical instabilities, i.e., numerical noises generate, accumulate, and finally explode. In our FDTD simulations we put several PML layers at z = −1.0 μm to shorten the z dimension of the computation domain in order to reach the necessary numerical precision. We conclude that the fine spectral features observed in Figure 3 are due to the photonic band structure of the porous membrane. Note that we were not able to simulate the real QD fluorescence peak, since by classical electrodynamics (FDTD) the wavelength width of the simulated light source must be wide enough so that the initial light will be spatially confined. Redshift in the Fluorescence Spectra. We now study the redshift observed in Figure 2b. One relevant issue about the combination of the QD and membrane-separation techniques is the possible requirement for high excitation power, since the number of QDs captured by the thin membrane can be limited. Figure 7a and b shows the spectral effects of varying the excitation power incident on a membrane with embedded QDs. The fluorescence peaks were fitted to Lorentz peaks and the resulting peak wavelengths are plotted against the excitation power in Figure 7c. A redshift is observed in the fluorescence spectra of QDs embedded in the membrane, while the wavelength of the fluorescence peak from QDs in dispersion remains the same, as seen in Figure 7d and ref 28. The redshift is most likely caused by phonons generated from electron−phonon interactions when the high-energy electron and hole, generated by the photon absorption, relax to the ground exciton state before the radiative recombination,
i.e., fluorescence.29 Phonons are accumulated in the QDs embedded in the membrane, heating up the QDs (immersed in immersion oil whose thermal diffusivity is very low, see below) thereby leading to the narrowing of the QD bandgap. In contrast, the heat is quickly dissipated from QDs into the continuous phase when QDs are in dispersion (sample B). Moreover, QDs embedded in the membrane are physically adsorbed to the membrane and thus not mobile, while QDs in dispersion are free to move and thus can shed heat more easily. Assume that an amount of heat is generated at a point-like QD located at r = 0, which increases the QD temperature to T0. The heat will diffuse into the surrounding medium according to the following equation ∂T (r, t ) = α∇2 T (r, t ) ∂t
(1)
with the initial boundary condition T(r, 0) = (T0 − T1) δ(r) + T1, where α and T1 are the thermal diffusivity and initial temperature of the surrounding medium, respectively. The solution to this problem is T (r , t ) =
T0 − T1 3/2
(4παt )
e −r
2
/4αt
+ T1
(2)
which shows that the temporal change of the QD temperature is determined by α. A large thermal diffusivity of the surrounding medium implies a fast change in the QD temperature. To estimate α of water and immersion oil, we performed the following measurement. 450 mL of water in a 500 mL glass beaker was heated on a hot plate with a magnetic stir bar to a stable and uniform temperature of 64.4 °C (continuously monitored). A two-neck glass flask was immersed into the heated water, with a thermometer plugged into one neck. Fifteen milliliters of water was put into the flask through the other neck, and the water temperature was recorded at a time interval of 10 s. The same volume of immersion oil was measured in the same way. The results are shown in Figure 8. Numerical simulations of eq 1, with boundary conditions of T(r > r0, t) = T1 and T(r ≤ r0, t = 0) = T0, where r0 is the radius of the glass flask, shows that the difference between water and oil 14154
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in Figure 8 is well fitted when αwater = 4αoil. Equation 2 thus indicates that the heat generated in the QD is well preserved by the immersion oil. Furthermore, we monitored the temperature of the membrane (sample A) under a continuous 408 nm excitation (15 mW) for 2 h and found no significant temperature change. This is expected since the optical bandgap of the aluminum oxide is more than 7 eV, much higher than the photon energy of 408 nm (3.04 eV, also of the 488 nm/2.54 eV laser in the confocal microscope). Finally, we heated up our dry QDs from 22 °C (room temperature) to 52 °C, measured the fluorescence spectrum under very weak excitation, and found a redshift of 4 nm. The fluorescence peak blueshifted back when the QDs cooled down to 22 °C. Such a temperature dependence agrees well with the work of Jing et al.30 This heating effect thus explains the redshift observed in Figure 2b.
SUMMARY
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AUTHOR INFORMATION
In a brief summary, we studied the fluorescence properties of colloidal CdSe-CsS/ZnS core-multishell QDs embedded in an inorganic porous membrane. Fine spectral features appeared superimposed on the principal QD fluorescence peak. FDTD simulations and further characterization of dispersed QDs revealed that the observed fine spectral features were caused by the photonic band structure of the porous membrane. Moreover, the principal fluorescence peak of QDs embedded in the membrane red-shifted when the excitation power was high, while it remained unchanged when QDs were in dispersion. The redshift was understood in terms of the QD material being heated by emitted phonons when the photoexcited electron and hole relax nonradiatively from high-energy states to the ground exciton state before fluorescence.
Corresponding Author
*E-mail: [email protected]. Notes
The authors declare no competing financial interest.
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REFERENCES
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Figure 8. Thermal diffusivity measurements of water and immersion oil.
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ACKNOWLEDGMENTS
The work was supported by the Swedish Research Council (621-2011-4381). H. X. acknowledges a scholarship from the China Scholarship Council and K. B. G. funding by the Swedish Research Council (B0460801). 14155
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Colloidal CdSe/CdS Quantum Dots. Phys. Chem. Chem. Phys. 2011, 13, 5848−5854. (24) http://rsbweb.nih.gov/ij/. (25) www.filmetrics.com/refractive-index-database/Al2O3. (26) Ruan, Z. C.; Qiu, M. Enhanced Transmission through Periodic Arrays of Subwavelength Holes: The Role of Localized Waveguide Resonances. Phys. Rev. Lett. 2006, 96, 233901. (27) Ding, B. Y.; Hrelescu, C.; Arnold, N.; Isic, G.; Klar, T. A. Spectral and Directional Reshaping of Fluorescence in Large Area SelfAssembled Plasmonic-Photonic Crystals. Nano Lett. 2013, 13, 378− 386. (28) Molnár, M.; Fu, Y.; Friberg, P.; Chen, Y. Optical Characterization of Colloidal CdSe Quantum Dots in Endothelial Progenitor Cells. J. Nanobiotechnol. 2010, 8, 2. (29) Fu, Y.; Ågren, H.; Kowalewski, J. M; Brismar, H.; Wu, J.; Yue, Y.; Dai, N.; Thylén, L. Radiative and Nonradiative Recombination of Photoexcited Excitons in Multi-Shell-Coated CdSe/CdS/ZnS Quantum Dots. Europhys. Lett. 2009, 86, 37003. (30) Jing, P. T.; Zheng, J. J.; Ikezawa, M.; Liu, X. Y.; Lv, S. Z.; Kong, X. G.; Zhao, J. L.; Masumoto, Y. Temperature-Dependent Photoluminescence of CdSe-Core CdS/CdZnS/ZnS-Multishell Quantum Dots. J. Phys. Chem. C 2009, 113, 13545−13550.
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Modulated Fluorescence of Colloidal Quantum Dots Embedded in a Porous Alumina Membrane Hao Xu,† Li Li,† Otto Manneberg,† Aman Russom,† Kristinn B. Gylfason,‡ Hjalmar Brismar,† and Ying Fu*,† †
Science for Life Laboratory, Department of Applied Physics, Royal Institute of Technology, SE-106 91 Stockholm, Sweden Micro and Nanosystems, Royal Institute of Technology, SE-100 44, Stockholm, Sweden
‡
ABSTRACT: The fluorescence spectrum of CdSe core-CdS/ZnS shell colloidal quantum dots (QDs) embedded in porous alumina membrane was studied. Small peaks, superimposed on the principal QD fluorescence spectrum, were observed. Finite-difference time-domain simulation indicates that the QD point radiation emitting from within the membrane is strongly modulated by the photonic band structure introduced by the membrane pores, leading to the observed fine spectral features. Moreover, the principal QD fluorescence peak red-shifted when the optical excitation power was increased, which is attributed to QD material heating due to emitted phonons when the photoexcited electron and hole relax nonradiatively from high-energy states to the ground exciton state before fluorescence.
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from Whatman Ltd.,21 as a first step toward the QD-membrane technique. In contrast to nanoporous membranes, the Whatman membranes show basically no optical response (see below). However, porous materials show extraordinary light reflection and transmission due to their photonic band structure.22 We therefore expect modulation of the QD fluorescence by the photonic band structure of the Whatman porous membrane, when QDs are embedded in the membrane, since the wavelength of the QD fluorescence is comparable with the pore size of the membrane. Nanoporous membranes, on the other hand, are not expected to affect the QD fluorescence in the visible range because of the nanopore sizes.
INTRODUCTION Colloidal quantum dots (QDs) have been attracting more and more researchers in the biomedical imaging field in the past decade. 1−4 Compared to traditional organic dyes and fluorescent proteins, QDs of nano size (2−10 nm) show great application potential due to their unique optical properties,5−7 such as high photoluminescence quantum yields, narrow emission spectra, tunable energy bandgaps, simultaneous excitation of multiple fluorescent colors, and superior photostability (much less photobleaching). Moreover, QDs, especially water-dispersible QDs, have been successfully and stably conjugated to specific biological molecules,8−10 and further used for in vitro cell labeling and in vivo imaging purposes.11,12 On the other hand, porous alumina membranes, with high pore density, tunable pore size and distribution, low protein binding, and minimal autofluorescence, are extensively used as biomolecular filters13 and for separating particles in liquids,14 for, e.g., proteomics and disease diagnostics.15−18 A potential application could be combining QDs and the membrane separation technique into one composite system, where selectively QD-conjugated biomolecules are captured in a porous membrane, through the attachment of specific surface ligands around QDs. This would enable rapid identification and in vitro diagnosis, through the simple monitoring of the QD fluorescence signal. Fluorescence spectra of QDs, e.g., CdSe/ZnS QDs19 and Si QDs,20 have been studied and reported after embedding QDs into nanoporous anodic alumina membranes, showing an unmodulated QD fluorescence spectrum and a modulated fluorescence from the nanoporous alumina membranes. Nanoporous (pore diameter of 50 nm) alumina membranes have a broad emission peak ranging from 350 to 600 nm. In this work, we study the fluorescence of QDs embedded in porous aluminum oxide membranes (Anodisc 47, 6809−5522) with pore size 0.2 μm and average thickness 60 μm purchased © 2013 American Chemical Society
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EXPERIMENTAL METHODS
Sample Preparation. Water-dispersible CdSe-CdS/ZnS coremultishell QDs with an emission-peak wavelength of 607 nm at room temperature were made in-house by common chemical synthesis (see our earlier work3,23). More specifically, a mixture of CdO (13 mg, 0.1 mmol), trioctylphosphine oxide (TOPO) (0.65 g, 1.7 mmol), ntetradecylphosphonic acid (TDPA) (56 mg, 0.2 mmol), and octadecylamine (ODA) (0.35 g, 1.3 mmol) was put into a 50 mL three-neck flask which was vacuumed for 30 min at 100 °C. After that, the flask was heated to 330 °C under nitrogen flow to get a clear solution. The temperature was then lowered down to 260 °C and 12 mg Se dissolved in 0.4 mL tributylphosphine (TOP) was injected quickly into the reaction mixture. We kept the temperature at 260 °C for the QDs core growth, then switched off the heat supply for the reaction flask when the desired QD size was reached. When the temperature came down to 100 °C, 15 mL acetone was added, leading to a precipitation of the QDs. Centrifugation and decantation were carried out subsequently several times to remove residual precursors and purify the QD cores. The following procedure was performed to add shell structures to the CdSe core: 1 g ODA, 4 mL 1-octadecene Received: September 12, 2013 Revised: October 16, 2013 Published: October 17, 2013 14151
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(ODE) were put into the 50 mL three-neck flask which was vacuumed for 30 min at 100 °C. The CdSe cores synthesized before were dissolved in chloroform and quickly injected into the reaction solution, which was then heated to and kept at 230 °C under nitrogen flow; Cd−, S−, and Zn− precursors were injected into the reaction mixture sequentially to get the desired core/shell CdSe QDs. Finally, 3mercaptopropionic acid (3-MPA) was used to replace ODA on the oildispersible QDs to make them water dispersible. The following samples were prepared and studied: Sample A: untreated membrane. The membrane was immersed in immersion oil (Immersol 518F, Zeiss) on a microscope slide and covered by a coverslip. Sample B: QDs in dispersion. QDs dispersed in an aqueous continuous phase at a concentration of 100 nM were dropped into a circular area formed by nail polish on a microscope slide, and covered by a coverslip. Excitation light was focused on the center of the circle. Sample C: membrane with embedded QDs. The membrane was immersed in a 3 nM QD dispersion for 100 min to adsorb QDs. It was then put into water for 20 min, to wash away loosely attached QDs, and subsequently air-dried and immersed into oil on a microscope slide. Sample D: highly diluted oil-soluble CdSe core/shell QDs (at a concentration of 10 pM) distributed on a microscope slide by dip coating, then dried at ambient temperature. The porous structure of a representative membrane was characterized by scanning electron microscopy (SEM), at an acceleration voltage of 15 kV, using an FEI Nova NanoSEM 650 system. Figure 1a shows a typical SEM image of the membrane surface and Figure 1b a typical cross-section, confirming membrane specifications.
Figure 2. (a) Optical reflectance spectra from the microscope slide (dashed line), and the sample A membrane on a slide (solid line), under 488 nm laser excitation. (b) Fluorescence spectra of the sample B QD dispersion (dashed line, peak position 607 nm), and the sample C membrane with embedded QDs (solid line, peak position 610 nm), from the circular area of diameter of 1.4 μm indicated in (c), with different normalization factors for direct comparison.
membrane with embedded QDs (sample C, circular collection area with a diameter of 1.4 μm, as indicated in Figure 2c), with different normalization factors for the convenience of comparison, showing that when the fluorescence spectrum of the embedded QDs is collected from a large area, it is very similar to that of QDs in dispersion. However, a small redshift was observed for the embedded QDs, which will be discussed below. Figure 3a shows the fluorescence spectra obtained from a series of circular areas of diameter 0.21 μm on sample C (see inset). Small peaks, superimposed on the principal QD fluorescence spectra, were observed. Figure 3b demonstrates how the small peaks (fine spectral features) are averaged out when the observation area is increased gradually (diameter increasing from 0.14 to 1.4 μm). To understand the fine spectral features in Figure 3a and b, we dispersed highly diluted oil-soluble CdSe core/shell QDs (10 pM) on a microscope slide (sample D). The QDs were so diluted that blinking was observed under continuous wave excitation (for experimental details, see ref 3). As shown in Figure 4a, we chose four single QDs marked by small rings in Figure 4c and measured their florescence spectra from circular areas with the same diameter of 0.21 μm, i.e., the same circular size as in Figure 3a. We further obtained another four fluorescence spectra (see Figure 4b) from neighboring QDs enclosed in large ellipses in Figure 4c. It is easily observed that the measured fluorescence spectra of various QDs on the structure-less microscope slide surface (Sample D) are rather smooth, much alike the QDs in dispersion. Moreover, the observed dispersion of the peak positions of single QD spectra in Figure 4a is only around 10 nm, and the fluorescence peak in Figure 4b from two neighboring QDs splits into two peaks with a distance of approximately 6 nm, while the fine spectral features in Figure 3 extend over a spectral width of at least 70 nm. It is therefore concluded that the fine spectral features presented in Figure 3 should be attributed to the porous structure of the membrane. Finite-Difference Time-Domain Simulation. We employed the finite-difference time-domain (FDTD) method to study the fine spectral features in the fluorescence from QDs embedded in porous membranes. The porous structure of the
Figure 1. Typical SEM images of the Whatman aluminum oxide membrane. (a) The surface morphology shows that the diameter of the pores is about 0.2 μm. (b) A cross-section shows that the thickness of the membrane is about 55 μm. Characterization Method. All samples listed above were characterized using an LSM 780 confocal microscope (Carl Zeiss, Jena, Germany) with a Plan Apochromat 63 × 1.4 Oil DIC M27 objective, and a 32-channel GaAsP spectral detector, at an excitation wavelength of 488 nm. The optical power of the excitation laser was varied. The fluorescence emission from the samples was recorded over a bandwidth of 546−671 nm with a spectral step of 2.9 nm. Each imaging frame contains 128 × 128 pixels, and the pixel size is 70 × 70 nm2; thus, the imaging size is 9.0 × 9.0 μm2. The software ImageJ24 was used to read the fluorescence signals from the images acquired by the microscope. The mean value of the pixels in the selected area was used for analysis.
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RESULTS AND DISCUSSION Fine Spectral Features in the Fluorescence Spectra. We first measured the optical response of the microscope slide and the membrane. Figure 2a shows the reflectance spectra under 488 nm illumination from a clean microscope slide and a clean membrane on a slide (sample A), indicating negligible optical response from the membrane. Figure 2b presents fluorescence spectra of the QD dispersion (sample B) and a 14152
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Figure 5. (a) Geometry of the FDTD simulation. The annotated x, y, and r indicate the positions and radii of the pores. (b) Optical spectra of the light recorded by the planar detector. Red dashed line: the spectrum of the point source in immersion oil; solid lines: spectra with immersed porous-membrane structure, when the point source is situated at different positions (shown by value of d corresponding to the left structure).
from z = −0.6 to z = 0.1 μm, i.e., from inside the membrane to outside the membrane. The simulation space along the z direction was between z = −1 and z = 3 μm, terminated by perfectly matched layers (PMLs). A planar detector was placed at z = 2.2 μm, to record the electric field which consists of the direct light from the point source as well as the reflected light from the membrane, the same geometric configuration as that of sample C. The mesh size of the FDTD simulations was 6 nm. The simulated optical spectra are presented in Figure 5b. The detected light intensity from a point source located at d = 0.1 μm was almost twice that of a source located inside the membrane, i.e., at d ∈ (0.0, −0.6) μm. The higher intensity is due to the reflection from the membrane. Weak ripples were seen in the spectrum of d = 0.1 μm, and also appeared in the point source because of the finite wavelength range involved in the numerical FDTD calculations. Figure 5 shows that the light from the point source was strongly modulated when the source was positioned on the membrane surface and inside the membrane, i.e., at d ∈ (−0.6, 0.0) μm, resulting in fine spectral features similar to those observed experimentally in Figure 3. Similar fine spectral features, but at different wavelengths, were obtained in simulated spectra, when the positions and sizes of the four pores in the membrane were varied. This explains the disappearance of the fine spectral features in Figure 3b when the observation area was enlarged. The photonic band structure modulation is only observable when the QD is positioned on the membrane surface or inside a pore of the membrane. The effects are clearly observed in the spatial distributions in Figure 6 of the electric fields from QDs outside and inside the pore, respectively, at two different times (time step 200 and 400, corresponding to real times of 220 and 440 ps, respectively). In the FDTD computation domain, the xy plane is periodic so that in Figure 6a we observe not only the electric field from the QD at x = 0.15 μm but also the electric field from the QD at x = 0.75 μm at t = 200, while in Figure 6b and d we observe the interference between the electric fields from the two QDs. The refractive index of the membrane is relatively low, and thus the degree of the spatial variation in the refractive index is limited along the z direction, so that the light from the QD
Figure 3. Fluorescence spectra features of QDs embedded in a membrane (sample C). (a) Fluorescence spectra obtained from a series of circular areas of diameter 0.21 μm along the vertical direction (arrow). (b) Fluorescence spectra from concentric circular areas with diameters increasing from 0.14 to 1.4 μm.
Figure 4. (a) Fluorescence spectra of the single oil-soluble CdSe core/ shell QDs indicated by small rings in (c). (b) Fluorescence spectra of neighboring QDs enclosed by large ellipses in (c). (c) One image frame showing the spatial locations of the QDs whose fluorescence spectra are presented in (a) and (b).
membrane was optically represented by the spatial variation of the refractive index, i.e., n = 1.5 in porous areas filled with immersion oil and n = 1.7 in the alumina.25 Figure 5a shows the geometric structure of our FDTD simulations. The membrane structure was simulated by a film of 0.6 × 0.6 μm2 periodically extended in the xy plane, with a thickness of 0.6 μm (z ∈ (−0.6, 0.0) μm). This membrane patch was irregularly perforated with four pores of diameters around 0.2 μm. The QD fluorescence was simulated by a point Gaussian-pulse source with a wavelength range from 540 to 690 nm centered at 607 nm, as illustrated by the red dashed line in Figure 5b. The point source was positioned at (0.15, 0.15, d) μm, where d was varied 14153
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Figure 7. Fluorescence spectra vs excitation powers (from power 20% to 100% at a step of 10%) from sample C and QD dispersion (1/ 10000, 1/3600, and 1/4000 are the intensity scaling factors applied). Fluorescence spectra under excitations of different laser powers (a) from a large area enclosing 650 pixels (3.18 μm2); (b) from a small area enclosing 140 pixels (0.69 μm2). (c) Peak wavelength vs excitation power for large (solid stars) and small (hollow stars) areas. (d) Spectra from QD dispersion (sample B) under different power excitations 20%, 50%, and 100%.
Figure 6. Electric field distributions at time step 200 (a,c) and 400 (b,d). (a,b) The QD (marked as “+”) is positioned at d = 0.2 μm; (c,d) the QD (marked as “+”) is positioned at d = −0.3 μm. Dashed lines mark the boundaries of the simulated membrane and the edges of the pore in the membrane.
outside the pore is largely reflected (see Figure 6a,b). The short-range ordered pores in the xy plane modify the light propagation in that plane and induce photonic resonance modes,26,27 which are clearly observed in Figure 6b,d. Note that the edges of the point light source look cubic in Figure 6a which is caused by the cubic FDTD lattice. Moreover, the membrane thickness in the FDTD simulation is only 0.6 μm, while it is about 55 μm in reality. As seen in Figure 6, the photonic resonance modes appearing in the electric field distribution are largely caused by the refractive index modulation in the xy plane, and the light reflection from the membrane boundary at z = −0.6 μm is very limited. A thin membrane is therefore necessary to be simulated. On the other hand, FDTD can, in theory, handle the 55-μm-thick membrane by extending the computation time. However, a long-time calculation can easily result in numerical instabilities, i.e., numerical noises generate, accumulate, and finally explode. In our FDTD simulations we put several PML layers at z = −1.0 μm to shorten the z dimension of the computation domain in order to reach the necessary numerical precision. We conclude that the fine spectral features observed in Figure 3 are due to the photonic band structure of the porous membrane. Note that we were not able to simulate the real QD fluorescence peak, since by classical electrodynamics (FDTD) the wavelength width of the simulated light source must be wide enough so that the initial light will be spatially confined. Redshift in the Fluorescence Spectra. We now study the redshift observed in Figure 2b. One relevant issue about the combination of the QD and membrane-separation techniques is the possible requirement for high excitation power, since the number of QDs captured by the thin membrane can be limited. Figure 7a and b shows the spectral effects of varying the excitation power incident on a membrane with embedded QDs. The fluorescence peaks were fitted to Lorentz peaks and the resulting peak wavelengths are plotted against the excitation power in Figure 7c. A redshift is observed in the fluorescence spectra of QDs embedded in the membrane, while the wavelength of the fluorescence peak from QDs in dispersion remains the same, as seen in Figure 7d and ref 28. The redshift is most likely caused by phonons generated from electron−phonon interactions when the high-energy electron and hole, generated by the photon absorption, relax to the ground exciton state before the radiative recombination,
i.e., fluorescence.29 Phonons are accumulated in the QDs embedded in the membrane, heating up the QDs (immersed in immersion oil whose thermal diffusivity is very low, see below) thereby leading to the narrowing of the QD bandgap. In contrast, the heat is quickly dissipated from QDs into the continuous phase when QDs are in dispersion (sample B). Moreover, QDs embedded in the membrane are physically adsorbed to the membrane and thus not mobile, while QDs in dispersion are free to move and thus can shed heat more easily. Assume that an amount of heat is generated at a point-like QD located at r = 0, which increases the QD temperature to T0. The heat will diffuse into the surrounding medium according to the following equation ∂T (r, t ) = α∇2 T (r, t ) ∂t
(1)
with the initial boundary condition T(r, 0) = (T0 − T1) δ(r) + T1, where α and T1 are the thermal diffusivity and initial temperature of the surrounding medium, respectively. The solution to this problem is T (r , t ) =
T0 − T1 3/2
(4παt )
e −r
2
/4αt
+ T1
(2)
which shows that the temporal change of the QD temperature is determined by α. A large thermal diffusivity of the surrounding medium implies a fast change in the QD temperature. To estimate α of water and immersion oil, we performed the following measurement. 450 mL of water in a 500 mL glass beaker was heated on a hot plate with a magnetic stir bar to a stable and uniform temperature of 64.4 °C (continuously monitored). A two-neck glass flask was immersed into the heated water, with a thermometer plugged into one neck. Fifteen milliliters of water was put into the flask through the other neck, and the water temperature was recorded at a time interval of 10 s. The same volume of immersion oil was measured in the same way. The results are shown in Figure 8. Numerical simulations of eq 1, with boundary conditions of T(r > r0, t) = T1 and T(r ≤ r0, t = 0) = T0, where r0 is the radius of the glass flask, shows that the difference between water and oil 14154
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in Figure 8 is well fitted when αwater = 4αoil. Equation 2 thus indicates that the heat generated in the QD is well preserved by the immersion oil. Furthermore, we monitored the temperature of the membrane (sample A) under a continuous 408 nm excitation (15 mW) for 2 h and found no significant temperature change. This is expected since the optical bandgap of the aluminum oxide is more than 7 eV, much higher than the photon energy of 408 nm (3.04 eV, also of the 488 nm/2.54 eV laser in the confocal microscope). Finally, we heated up our dry QDs from 22 °C (room temperature) to 52 °C, measured the fluorescence spectrum under very weak excitation, and found a redshift of 4 nm. The fluorescence peak blueshifted back when the QDs cooled down to 22 °C. Such a temperature dependence agrees well with the work of Jing et al.30 This heating effect thus explains the redshift observed in Figure 2b.
SUMMARY
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AUTHOR INFORMATION
In a brief summary, we studied the fluorescence properties of colloidal CdSe-CsS/ZnS core-multishell QDs embedded in an inorganic porous membrane. Fine spectral features appeared superimposed on the principal QD fluorescence peak. FDTD simulations and further characterization of dispersed QDs revealed that the observed fine spectral features were caused by the photonic band structure of the porous membrane. Moreover, the principal fluorescence peak of QDs embedded in the membrane red-shifted when the excitation power was high, while it remained unchanged when QDs were in dispersion. The redshift was understood in terms of the QD material being heated by emitted phonons when the photoexcited electron and hole relax nonradiatively from high-energy states to the ground exciton state before fluorescence.
Corresponding Author
*E-mail: [email protected]. Notes
The authors declare no competing financial interest.
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REFERENCES
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Figure 8. Thermal diffusivity measurements of water and immersion oil.
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ACKNOWLEDGMENTS
The work was supported by the Swedish Research Council (621-2011-4381). H. X. acknowledges a scholarship from the China Scholarship Council and K. B. G. funding by the Swedish Research Council (B0460801). 14155
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