Communication pubs.acs.org/cm
Synthesis of Robust Sandwich-Like SiO2@CdTe@SiO2 Fluorescent Nanoparticles for Cellular Imaging Yian Zhu,† Zhen Li,†,* Min Chen,‡ Helen M. Cooper,‡ Gao Qing (Max) Lu,† and Zhi Ping Xu†,* †
ARC Centre of Excellence for Functional Nanomaterials, Australian Institute for Bioengineering and Nanotechnology, and The Queensland Brain Institute, The University of Queensland, Queensland 4072, Australia
‡
S Supporting Information *
KEYWORDS: CdTe quantum dots, silica coating, reverse microemulsion, nanoparticles, cellular imaging
I
n the past decade, semiconductor quantum dots (QDs) have shown great potential in biolabeling and bioimaging1−4 due to their exceptional optical properties including size-tunable and narrow emission, wide absorption, and high photostability.5 However, the practical applications of QDs are hindered by several problems such as photo and chemical instability in harsh environments and cytotoxicity caused by the release of heavy metal ions.6,7 To resolve these problems, coating QDs with biocompatible silica is a promising alternative.8−11 Encapsulating QDs within silica shells has great advantages, since a silica shell can not only protect QDs from oxidation but also enable further surface functionalization with diverse groups. Recently a number of silica-coating procedures have been developed, mainly classified into the Stöber method12−14 and reverse microemulsion approach.8,15 Both methods, however, suffer from a huge drop of QD fluorescence during encapsulation.16−19 In some cases, the fluorescence of QDs has been completely quenched, which make QDs useless in labeling and imaging. Many efforts have been made to retain the fluorescence, such as reducing the electrostatic repulsion between QDs and silica intermediates,20 coating the QDs core with a wide-gap semiconductor shell,21 or incubating QDs in alkaline solution prior to silica coating.22 The highest fluorescence quantum yield (QY) of QD-silica nanoparticles (NPs) reported up to now is < 50%.22−24 It should be noted that most investigations are focused on preparation of QD-silica NPs, while there are only few reports on their bioapplication and cytotoxicity due to the poor fluorescence QY.25 This communication reports a new approach to prepare robust QD-silica NPs with higher fluorescence QY and lower cytotoxicity. As briefly outlined in Scheme 1, the synthesis
Figure 1. (A) TEM image of thiol-capped silica NPs. (B) PL spectra of thiol-capped silica before and after isoindole conjugation. The inset shows the photographs of silica before (left) and after (right) conjugation excited with 365 nm UV light. (C) XPS spectra of thiolcapped silica NPs.
of the surface −SH group was confirmed by reacting with o-phthaldialdehyde and glycine to generate fluorescent 1-alkyithio-2-alkyl-substituted isoindole (Supporting Information Scheme S1).27 As shown in Figure 1B, the strong photoluminescence (PL) at 530 nm suggests the successful graft of −SH groups onto the surface of silica NPs. The presence of −SH groups was further proved by the XPS analysis. In the overview spectrum (Supporting Information Figure S2), the binding energies of O 1s, S 2p, Si 2s, and Si 2p were found to be 532.8, 163.5, 154.2, and 103.4 eV, respectively. The binding energy profile of S 2p was shown in Figure 1C. The spectrum was fitted by two peaks located at 163.6 and 164.8 eV, attributed to S 2p3/2 and S 2p1/2 (Supporting Information Figure S3 and Table S1).28 The sulfur content was determined to be 1.06 wt % by elemental analysis, and the density was calculated to be ∼4 −SH/nm2. The grafted thiol groups show strong affinity to CdTe nanocrystals (NCs) which in situ grow on the silica surface to form core−satellite NPs. In order to increase the fluorescence
Scheme 1. Illustration of the Procedure for Preparation of SiO2@CdTe@SiO2 Nanoparticles
starts from the thiol-capped silica nanospheres.26 Figure 1A shows the TEM image of silica NPs with a diameter of 32.7 ± 2.5 nm (Supporting Information Figure S1A). The existence © 2012 American Chemical Society
Received: November 10, 2011 Revised: January 12, 2012 Published: January 23, 2012 421
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of the NPs, mercaptosuccinic acid (MSA) was used as a costabilizer during preparation. After reaction, CdTe NCs can be clearly seen to reside on the surface of silica NPs (Figure 2A).
Figure 3. (A−C) TEM and HRTEM images of SiO2@CdTe@SiO2 NPs. (D) PL spectra of SiO2@CdTe and SiO2@CdTe@SiO2 NPs. The inset is the photographs of the SiO2@CdTe@SiO2 suspension under room light and UV light of 365 nm.
Figure 2. (A) TEM image, (B) UV/vis absorption and PL spectra, and (C) XPS spectra of core−satellite SiO2@CdTe NPs.
The average size of SiO2@CdTe NPs slightly increased to 36.4 ± 4.1 nm (Supporting Information Figure S1B). Figure 2B displays the UV/Vis absorption and PL spectra of the SiO2@CdTe NPs, showing that by adjusting the refluxing time, a set of fluorescent NPs with different emission peaks can be synthesized (Supporting Information Figure S4). Determination of the amounts of cadmium and tellurium in particles and in the supernatant (Supporting Information Table S2) by inductively coupled plasma optical emission spectrometer (ICP-OES) reveals that as-generated CdTe NCs were mostly deposited on silica beads and show high fluorescence (Supporting Information Figure S5), largely due to the presence of surface −SH groups. The fluorescence QYs of three as-prepared SiO2@ CdTe NPs were 31.8%, 52.3%, and 72%, respectively. Similarly, the XPS results (Supporting Information Figure S2) indicate the presence of Si, O, C, S, Cd, and Te elements. The characteristic binding energies of Cd 3d5/2 and Te 3d5/2 are located at 405 and 572 eV (Figure 2C), respectively, consistent with those of CdTe QDs reported elsewhere.29,30 The similar binding energy of S 2p found in thiol-capped silica and SiO2@CdTe NPs (Supporting Information Figure S3) indicates the similar chemical environment of thiol groups. In order to retain the fluorescence of CdTe QDs and increase their stability, another silica layer was then coated onto the surface of the SiO2@CdTe NPs via the reverse microemulsion method. Here the SiO2@CdTe aqueous solution was incubated with MPS prior to coating, which further helped protect the QDs and resulted in a high retention of the fluorescence. As shown in Figure 3A, the average size of NPs increased to 67.5 ± 6.7 nm (Supporting Information Figure S1C) after coating, indicating that the thickness of the silica layer was ∼15 nm. The TEM image presented in Figure 3B clearly shows the sandwich structure of SiO2@CdTe@SiO2 NPs. The HRTEM image (Figure 3C) shows an average interplanar spacing of 0.19 nm, corresponding to the lattice of CdTe NC and indicating that CdTe QDs were well-crystallized. Figure 3D displays the PL curves of SiO2@CdTe (refluxing for 24 h) and corresponding SiO2@CdTe@SiO2 NPs. By comparing their
integrated areas, we concluded that 85% of the initial fluorescence was retained. The fluorescence QY of SiO2@ CdTe@SiO2 was ∼61%, the highest value yet achieved to the best of our knowledge. In addition, a slight blue-shift of the fluorescence peak was observed, probably due to the etching of CdTe QDs caused by ammonia.16 According to the XPS analysis results (Supporting Information Figure S2), no signals of Cd, Te, or S were detected in the SiO2@CdTe@SiO2 sample, indicating that CdTe NCs were completely coated as shown in Figure 3B. The successful silica coating is expected to improve the chemical stability of QDs. Supporting Information Figure S6 shows the fluorescence efficiency of CdTe, SiO2@CdTe, and SiO2@CdTe@ SiO2 NPs against different pH values. The fluorescence of CdTe QDs is strongly influenced by minor variations in solution pH, indicating poor chemical stability. In contrast, SiO2@CdTe NPs are stable in a pH range of 3−9. However, their fluorescence was markedly decreased under acidic and basic conditions. For SiO2@ CdTe@SiO2 NPs, the fluorescence was stable in a wider pH range (3−13) and only weakened at pH < 3. Therefore, SiO2@CdTe@ SiO2 NPs show a significantly improved chemical stability compared with CdTe QDs and SiO2@CdTe NPs. These findings strongly demonstrate that highly fluorescent and chemical stable SiO2@CdTe@SiO2 NPs have been successfully obtained through the newly designed procedure. The high fluorescence retention can be attributed to two factors. One is the thiol group capping on the core silica surface which allows the CdTe NCs to tightly adhere to the silica core. The other is the incubation of SiO2@CdTe NPs with MPS prior to the silica coating, which directs MPS to precoat and protect CdTe QDs and induces the subsequent compact silica coating. Without MPS involvement, the fluorescence of prepared SiO2@CdTe@SiO2 NPs was less than 20%. The synthesized SiO2@CdTe@SiO2 NPs are expected to show low cytotoxicity. Figure 4A,B displays the cell viability of the human embryonic kidney cell line (HEK 293 cells) in the presence of CdTe QDs and SiO2@CdTe@SiO2 NPs suspensions. At comparable concentrations of CdTe, only 70% of cells survived 422
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gratefully acknowledges the support from Queensland Smart Futures Fellowship, UQ early career-research grant, and UQ new staff research startup grant. H.M.C. gratefully acknowledges support from the Queensland Smart Futures Fellowship Scheme. Z.P.X. gratefully acknowledges the ARC Australian Research Fellow (ARF) Scheme.
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Figure 4. Cell viability of (A) MSA-capped CdTe NCs and (B) SiO2@ CdTe@SiO2 in HEK293 cells and (C) CLSM images of HEK 293 cells after incubation with SiO2@CdTe@SiO2 for 2 h: (left) QDs, (middle) DAPI, (right) overlay of QDs and DAPI. Scale bar: 20 μm.
after incubation with CdTe QDs for 4 h and then being cultured for 48 h. In contrast, SiO2@CdTe@SiO2 NPs exhibited much lower cytotoxicity than CdTe NCs (greater than 90% cell viability) and were slightly affected at higher concentrations (75 μg/mL). The confocal laser scanning microscopy (CLSM) image (Figure 4C, left) shows that the yellow SiO2@CdTe@SiO2 NPs (PL peak: 585 nm) can be seen in both the cytoplasm and the nucleus of HEK 293 cells after a 2 h incubation, indicating that the sandwichlike NPs were efficiently internalized by the cells, even without any other transfection technique. These data suggest the potential of these SiO2@CdTe@SiO2 NPs as robust biomarkers due to their stronger fluorescence and lower cytotoxity. In conclusion, we have developed a versatile process for the synthesis of highly fluorescent and monodispersed SiO2@ CdTe@SiO2 sandwich-structured NPs with an average diameter of less than 100 nm. As-prepared SiO2@CdTe@SiO2 NPs displayed a higher fluorescence QY and lower cytotoxicity in comparison with their QDs-silica analogues. These advantageous properties and the ability for further functionalization using straightforward chemistry make SiO2@CdTe@SiO2 NPs very attractive for biomedical imaging applications.
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ASSOCIATED CONTENT
S Supporting Information *
Experimental details and characterization data (PDF). This material is available free of charge via the Internet at http:// pubs.acs.org.
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REFERENCES
AUTHOR INFORMATION
Corresponding Author
*Tel.: +61-7-3346 3817. Fax: +61-7-3346 3973. E-mail: z.li3@ uq.edu.au (Z.L.);
[email protected] (Z.P.X.). Notes
The authors declare no competing financial interest.
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ACKNOWLEDGMENTS This work has been financially supported by the Australian Research Council through a Discovery Project Grant (DP0879769). Y.Z. thanks the UQ International Research Tuition Awards and Gregg Thompson scholarship. Z.L. 423
dx.doi.org/10.1021/cm2033417 | Chem. Mater. 2012, 24, 421−423
