Open Access
Add to BibSonomyAbstract
Indocyanine green-loaded mesoporous silica-coated gold nanorods (ICG-loaded Au@SiO2) were prepared for the dual capability of X-ray computed tomography (CT) and fluorescence imaging. X-ray CT scanning showed that ICG-loaded Au@SiO2 could provide significant contrast enhancement; Near-infrared fluorescence generated by the nanomaterial was present up to 12 h post intratumoral injection, thus enabling ICG-loaded Au@SiO2 to be used as a promising dual mode imaging contrast agent. Multiplexed images can be more easily obtained with this novel type of multimodal nanostructure compared with traditional contrast agents. The dual mode imaging probe has great potential for use in applications such as cancer targeting, molecular imaging in combination with radiotherapy, and photothermolysis.
©2011 Optical Society of America
1. Introduction
Technical innovations that are economical and practical continue to maximize the ability of current imaging tools. One of the most feasible and effective innovation is a complementary contrast agent or imaging probe that is introduced to improve signal detection and sensitivity. The development of contrast agents and imaging probes forms an essential research field in biological and medical imaging and enhances the analysis of biological information, clinical diagnosis, and analysis of functional alterations at the cellular level.
X-ray computed tomography (CT) is one of the most widely used diagnostic tools in terms of frequency of use and cost. Nevertheless, there have been very few fundamental improvements in clinical CT contrast agents over the last 25 years. Hard tissues have higher X-ray attenuations than various soft tissues, whereas the contrast between soft tissues is inherently poor and this limits diagnostic sensitivity when investigating pathologies such as cancer. A major challenge in X-ray imaging of biological systems is the realization of a high local concentration of the contrast agent and a signal that surpasses the X-ray absorption of oxygen and carbon, the two major background components. Current contrast agents for CT are based on small iodinated molecules that only allow very short imaging times due to rapid clearance by the kidney, and may also cause renal toxicity [1,2]. Recent advances in nanotechnology have led to the development of novel types of contrast agents because of the uniquely useful electronic, optical, and magnetic properties of nanomaterials [3–7].
Gold has a higher atomic number and absorption coefficient than commercially available iodine-based contrast agents [8,9] and achieves better contrast and less bone and tissue interference with a lower X-ray dose. In addition, gold is non-toxic, has low osmolality and viscosity even at very high concentrations, unlike viscous iodine agents, and helps avoid side-effects. Previously, gold nanoparticles (AuNPs) have been applied to X-ray CT [10–16]. Properly treated AuNPs have a longer circulation time than iodine-based molecular contrast agents, which makes clinical observation and disease diagnosis more convenient and patient-friendly [11]. More importantly, it has been demonstrated that gold nanoparticles conjugated to biological directing agents, for example peptides or antibodies that enable visualization of disease-specific biomarkers at the molecular levels, are suitable for use as CT contrast agents [17–19]. It is also worth noting that if tumors can be loaded with gold this will lead to higher X-ray absorption by the cancerous tissue compared with normal tissue; higher tumor X-ray absorption due to AuNPs greatly improves the efficacy of radiotherapy treatment [20,21].
Nano-carriers containing near-infrared (NIR) exogenous chromophores are currently under investigation for use as site specific and sensitive optical imaging probes. This is the result of minimizing intrinsic background interference, relatively deep optical penetration, and substantial improvement in molecular and morphological contrast [22,23]. Recently, gold nanorods (GNRs) have been incorporated with the imaging moiety of the NIR fluorescent molecule, indocyanine green (ICG, a tricarbocyanine dye approved by United States Food and Drug Administration), via covalent or ionic bonding. This conjugate has been used for optical imaging and photodynamic therapy [24–26]. The most exciting feature of nanoparticle usage in biomedicine is the possibility that several payloads/features can be included to enable multiple purpose applications, for example, synergistic diagnostics and therapy, or multimodal imaging [19,27–29]. The development of extensive multifunctional and multimodal nanostructures achieved by packing together attractive properties, such as biomarkers, magnetism, and fluorescence, into a single nano-object will allow clinicians to obtain comprehensive morphological and molecular profiles for accurate clinical diagnosis [30,31].
AuNP-based CT contrast agents have been widely reported in the past [10–18], and now GNRs are beginning to attract considerable attention. They are being exploited in CT imaging due to their unique properties in photothermolysis [32–35], biosensing [36], molecular imaging [37], and gene delivery in cancer therapy [38]. Although the X-ray attenuation coefficient of GNRs for molecularly targeted cells in vitro is over five times higher than for identical but untargeted cancer cells or for normal cells [17], recent innovations have not improved this outcome.
CT is an anatomical imaging technique for which the signal attenuation is proportional to agent concentration Optical techniques are highly sensitive, very fast and can visualize multiple species with exceptional soft tissue contrast, but can only be reliably applied to small organisms, such as mice, because of the limited penetration of light. Therefore, a nanoparticle probe that simultaneously enhances CT contrast and NIR optical imaging could potentially be very valuable for many practical applications. The combination of these two techniques can give quantifiable information of contrast agent accumulation at different levels, which may prove especially valuable in a preclinical setting.
We designed a dual mode contrast agent for X-ray CT and NIR fluorescence imaging based on GNRs. We describe nanocomposite synthesis, quantitative contrast characterization, and present results of in vivo dual mode imaging. To the best of our knowledge, this is the first report on mesoporous Au@SiO2 loaded with an organic NIR dye as a dual mode imaging probe.
2. Experimental details
2.1 Reagents
Cetyltrimethylammonium bromide (CTAB), HAu(III)Cl4, NaBH4, AgNO3, L-ascorbic acid, NH4OH (25.2-28.0%) and anhydrous ethanol were purchased from Sinopharm Chemical Reagent Co., Ltd. (Shanghai, China). Tetraethoxysilane (TEOS) and 3-aminopropyltrimethoxy silane (APTES, >99%) were purchased from J & K Chemical Ltd. (Shanghai, China). Indocyanine green (ICG) (Tokyo Chemical Industry Co., Ltd., Tokyo, Japan) was dissolved in anhydrous ethanol and stored at 4°C in the dark (stock solution). Deionized water (Millipore Milli-Q grade) with a resistivity of 18.2 MΩ was used in all experiments.
2.2 Synthesis of GNRs and Au@SiO2
The GNRs were prepared according to the simple seed-mediated template-assisted protocol [39,40], by reducing gold salt in the presence of surfactant directed synthesis.
A 20 mL aliquot of GNRs stock solution was centrifuged and redispersed in 20 mL deionized water. Then 1.1 mL of the TEOS ethanol solution (10 mM) was added to the 20 mL of aqueous GNRs (pH adjusted to 10–11 by NH4OH). After vigorous stirring for 10 h at room temperature an approximately 13 nm thick silica layer formed on the surface of the GNRs through hydrolysis and condensation of TEOS [41]. The silica-coated nanoparticles were isolated by centrifugation, washed with deionized water and ethanol several times, and then dispersed in 20 mL of deionized water for later use.
2.3 ICG-loaded Au@SiO2
ICG was embedded into Au@SiO2, ICG by stirring it with APTES (ICG-APTES) in the presence of anhydrous ethanol [42]. Typically, 10 mg of ICG was added to 2.5 mL of anhydrous ethanol and stirred for 5 min before adding 100 μL of APTES. Twenty mL of the prepared Au@SiO2 was added to the appropriate amount of CTAB and 10 μL ICG-APTES, after which 0.6 mL of TEOS was added to mixture and the reaction allowed proceeding for another 12 h. The nanoparticles were collected by centrifugation, washed at least five times with ethanol and deionized water, and then dispersed in deionized water for later use.
2.4 Characterization
Samples were placed on carbon coated copper grids and analyzed with a transmission electron microscope (JEOL JEM-2100) operating at an accelerating voltage of 200 kV. UV–vis absorbance spectra were measured with a Shimadzu UV-2450 spectrophotometer. Fluorescence spectra were recorded on a HITACHIH FL-4600 spectrofluorimeter. Fourier transform infrared (FTIR) spectra with a wavenumber ranging from 400 to 4000 cm−1 were acquired using a Bruker EQUINOX 55 FTIR Spectrometer.
2.5 Animal model loaded with gastric cancer
All animal operations were in accordance with institutional animal use and care regulations. Female nude mice weighing 22 - 24 g were purchased from Shanghai SLAC Laboratory Animal Co., Ltd. (Shanghai, China). Five-week-old female nude mice were given a right rear flank subcutaneous (s.c.) injection of 5 × 106 MGC803 cancer cells in 100 mL of serum free RPMI-1640 medium. The MGC803 cell line was made available by the Chinese Academy of Science (Department of Bio-Nano Science and Engineering). Cell culture products and reagents were purchased from GIBCO.
3. Results and discussion
The deposition of a silica shell on the anisotropic GNRs was successfully completed using a modified Stǒber method [43]; the monodisperse Au@SiO2 structures had aspect ratios of 2.9 ± 2.7 (length: 44.44 ± 4.7 nm, width: 15.10 ± 1.7 nm) for the GNR cores and had a uniform silica shell thickness (~13 nm), as shown in Fig. 1 .
Fig. 1 TEM images of the prepared mesoporous Au@SiO2 with different scale bar. (a): 50 nm, (b): 20nm.
As seen in Fig. 2(a) , the prepared GNRs had a weak transverse plasmon band at 513 nm and a strong longitudinal plasmon band at 753 nm. The plasmon absorption band of the GNRs was chosen to avoid re-absorption of the emission by the NIR chromophores. After being coated with silica, there was an obvious 8 nm red-shift in the longitudinal peak compared with that of uncoated anisotropic GNRs but a negligible shift in the transverse plasmon band, which was the result of an increase in the local refractive index of the surrounding medium for GNRs after the formation of the silica shell by replacing CTAB [44]. Figure 2(b) shows that the ICG-loaded Au@SiO2 has absorption spectra at 396 nm, which corresponds to the absorption profile of ICG. In addition, only a small peak appears at 810 nm, and was likely due to fluorescence resonance energy transfer (FRET) [45] or suppression of the plasmon resonance [46]. Figure 2(c) represents the fluorescence emission spectra of ethanol dispersion of ICG and ICG-embedded into Au@SiO2, the encapsulation of ICG molecules Au@SiO2 similar emission wavelength compared with free form ICG solutions. Figure 2(d) was obtained using FTIR. The two peaks at 2915, 2855 cm-1 correspond to the asymmetric and symmetric stretching vibrations of the methylene group, respectively, which exists in the hydrolysate of APTES. The above data suggest that ICG was successfully incorporated into the nanocarriers. This demonstrates the synthesis of uniform-sized mesostructured silica-coated GNRs loaded with an organic cyanine dye using physical adsorption or electrostatic adsorption
Fig. 2 Characterization of prepared samples. The UV-vis absorbance spectra (a): uncoated GNRs and Au@SiO2, (b): ICG-loaded Au@SiO2 (Inset: absorbance profile of ICG). (c) Fluorescence emission spectra for ICG and ICG-loaded Au@SiO2 in ethanol; λex = 785 nm. (d) The FTIR spectra of ICG and ICG-loaded Au@SiO2.
A standard curve of CT image intensity versus Au@SiO2 concentration was plotted in Fig. 3(a) from a dilution series in which Au@SiO2 concentrations were determined before lyophilization and then dispersed in PBS (pH 7.4). Measurements were made using a clinical CT scanner (Siemens ONCOR impression OptiVue) at 130 keV and a slice thickness of 3 mm. A linear correlation was found between the Hounsfield units (HU) and Au@SiO2 concentration (R = 0.96225). X-ray absorption was directly proportional to the concentration of the contrast agent, such that HU values increased with higher Au@SiO2 concentrations (mg/mL). These results are in agreement with previous data [15], and indicate the potential usefulness for CT imaging.
Fig. 3 X-ray CT of Au@SiO2. (a) Concentration–signal curve obtained from the CT images of Au@SiO2 in PBS media (Inset: the corresponding Au@SiO2 concentrations are given in mg/mL); the HU value of PBS was 8. (b) CT images of a mouse viewed from the rear 1 min and 15 min after an intratumoral injection of Au@SiO2. Red circles indicate regions with enhanced contrast in the gastric tumor; the green arrow indicates a black hole at the needle injection site. Evaluation of Au@SiO2 contrast enhancement was carried out by loading digital CT images into a standard display program and then selecting a uniform round region of interest for each sample.
The feasibility of using ICG-loaded Au@SiO2 as an X-ray CT contrast agent was tested using xenograft mice with an approximately 5 mm diameter gastric cancer. A 200 μL (3 mg/mL) dose of Au@SiO2 was intratumorally administered and X-ray images of the tumor obtained 1 min (43 HU) and 15 min (56 HU) post injection, as shown in Fig. 3(b). Note that 30 HU is considered sufficient to distinguish between different tissues and corresponds to 5 mM Au [47]. The HU values reflect tissue density and are used to diagnose the disease. The enhanced local signal seen 15 min after the injection, increased local X-ray attenuation (56 HU) above normal soft-tissue values (~40 HU [48,49]), thus providing encouraging initial indications that sufficient specificity can be estimated in in vivo experiments.
The above results demonstrated that Au@SiO2 exhibited enhanced CT contrast. Some information about attenuation by the object that is carried by the transmitted X-ray flux is lost during the detection process. Hence, it is reasonable to ask to what extent the additional information acquired, using more advanced detection technology, can be used to obtain a better clinical diagnosis, a reduction in administered dosages, or to the development of versatile applications. We used ICG-loaded Au@SiO2 in combination with X-ray and NIR fluorescence imaging (In Vivo Imaging System, Lumina XR, Caliper Life Sciences, Inc., Hopkinton, MA01748, USA) to determine whether it could be used as an effective dual-mode imaging contrast agent.
Figures 4(a) and (c) show that it is difficult to discern differences between adjacent soft tissue and the tumor in planar projection X-ray images following a 15 s X-ray exposure. Increasing the exposure time to 30 s showed that enhanced contrast could be achieved following an intratumoral injection of ICG-loaded Au@SiO2 when compared with X-ray images using the same exposure time but without an injection (cf. Figures 4(b) and (d)). Nanocarrier concentration and its resultant gold content in the gastric cancer tissue was not significantly different 12 h after the injection when compared with the concentration seen at 0 and 6h, the spread of the nanoparticles following an intratumoral injection is not significant, thus the X-ray signals from the cancerous tissue had similar contrasts for up to 12h post injection (Fig. 5 ).
Fig. 4 The effects of exposure time on planar projection X-ray images. X-ray images using 15 s (a) and 30 s (b) exposure times before an ICG-loaded Au@SiO2 injection. X-ray images using 15 s (c) and 30 s (d) exposure times immediately after an injection of 200 μL of ICG-loaded Au@SiO2 (3 mg/mL). The green arrow indicates the tumor.
Fig. 5 In vivo series of planar X-ray images (30 s exposures) of an animal following an injection of 200 μL of ICG-loaded Au@SiO2 (3 mg/mL). The green arrow indicates the gastric cancer tumor.
Figure 6(a) shows planar projection X-ray images of different sample aliquots using the following operating parameters: current, 100 mA; tube voltage, 35 keV; exposure time, 3 s; fov, 10. The quantification of fluorescence intensity in Fig. 6(b) was recorded as radiant efficiency (), λex = 745 nm, λem = 790 nm using an exposure time of 1 s. Although there are no significant contrast differences between the X-ray images of the samples in Fig. 6(a) when view with the naked eye, sample I can be clearly distinguished from the others. Although these X-ray images are not easily differentiated visually (shown for the sake of completeness), a clinical CT scanner can quantitatively determine the HU values, and is much more sensitive, and has a higher spatial resolution [50] compared with the planar projection X-ray system. Additionally, the maximum voltage used for planar X-ray imaging was 35 keV, depending on the amount of filtration used. In contrast, CT scanning uses 130 keV. Gold imaging at 80 – 100 keV has a higher theoretical X-ray attenuation and contrast-to-noise ratio compared with lower keV values, thus this effectively reduces interference from bone absorption and takes advantage of lower soft-tissue absorption [49]. Fluorescence was only observed in samples III and IV in Fig. 6(b) when using epi-fluorescence, indicating the presence of the small molecular chromophore. It is well known that fluorescent activity is quenched when a fluorophore comes directly in contact with a metal surface due to nonradiative energy transfer from the excited state of the molecule to the metal. However, in our experiment the fluorescent chromophore in the original ICG suspension was added after the first injection of TEOS and contained within the silica shell using physical or electrostatic adsorption methods. Thus, there was a thin layer of silica sandwiched between the GNR and ICG chromophores that protected the ICGs from fluorescent quenching.
Fig. 6 ICG-loaded Au@SiO2 was examined using X-ray and NIR fluorescence dual mode imaging. Planar X-ray (a) and NIR fluorescence images (b) were obtained of 7 mg/mL Au@SiO2 (I), PBS (II), 3.2 mg/mL ICG-loaded Au@SiO2 (III) and 1.5 mg/mL ICG-loaded Au@SiO2 (IV). (c) In vivo planar X-ray images (exposure time 30 s) of a mouse prior to and 12 h post intratumoral injection of ICG-loaded Au@SiO2 (200 μL, 1.5 mg/mL). (d) An in vivo planar X-ray image using a 60 s exposure time (left) taken 12 h post intratumoral injection of the dual mode imaging contrast agent was overlapped with the homologous NIR fluorescence image (10 s exposure time) (right). Inset: the corresponding overlay of bright field and NIR fluorescent images; the quantification of fluorescence intensity was recorded as radiant (), with an exposure time of 10 s. The green arrow indicates the tumor. The images were reconstructed using the software supplied by the manufacturer.
The feasibility of using ICG-loaded Au@SiO2 as an in vivo contrast agent for dual imaging was demonstrated following a 200 μL, 1.5 mg/mL intratumoral injection. Planar X-ray in vivo tumor images (30 s exposures) before and 12 h after the injection show some post injection X-ray attenuation bright spots in Fig. 6(c). The corresponding NIR fluorescence of the ICG-loaded Au@SiO2 is clearly seen in the overlapped X-ray and NIR fluorescence images in Fig. 6(d). Intriguingly, when the planar X-ray exposure time was increased to 60 s, the subtle anatomy of the site and the bright spots seen in Fig. 6(c) were not obvious, so we inverted image and adjusted the levels. ICG is typically administered as an intravascular bolus injection and has a relatively short half-life in the blood (2-4 min) due to its rapid adsorption by albumin and high-density lipoproteins. The presence of the fluorescent ICG-loaded Au@SiO2 12 h post injection in Fig. 6(d) confirms the successful incorporation of ICG into the mesoporous silica. The encapsulation of the ICG within the silica after the second injection of TEOS protects the ICG against the external bioenvironment, and thus reduces non-specific protein binding and prolongs the vascular half-life of the ICG.
The sustained release effect of ICG-embedded in the mesoporous silica compartments was demonstrated using real-time in vivo NIR imaging of a mouse after subcutaneously injected into its chest of 200 μL of ICG-loaded Au@SiO2 (1.5 mg/mL) (Fig. 7 ). It is clear that after the injection fluorescence intensity gradually increased due to the release of ICG from the mesoporous silica shell.
Fig. 7 Real-time in vivo NIR images of a control (left) mouse and a mouse with a 200 μL subcutaneous chest injection of ICG-loaded Au@SiO2 (1.5 mg/mL).
Among the variety of inorganic materials studied for biomedical applications, silica-based structures have good biocompatibility, high tolerance against many organic solvents, and are thermally more stable under high-energy nanosecond laser irradiation [51]. Moreover, the surface of mesoporous silica structures can be easily functionalized for specifically targeting tumor cells in vivo. Mesoporous silica based nanoparticles are versatile frameworks that have advantages over other carriers due to their high pore volume and large surface area making it easier to incorporate various active molecules and metal nanocrystals [52,53]. Free ICG at concentrations of 10 μM show a significant decrease in fluorescence emission because ICG is known to self-quench in aqueous solutions at concentrations above 5 μM [54]. The exogenous ICG chromophores are mainly located in the mesoporous channels in ICG-loaded Au@SiO2 nanocomposites. The mesoporous silica shell provides spatially well-separated ICG molecules that can efficiently prevent them from aggregation and thus decrease self-quenching fluorescence. The growth of the second additional silica layer smoothed out the surface of the mesoporous silica and reduced the aspect ratio but it is not clear if the additional growth of silica completely filled the mesoporous holes of the initial silica layer.
4. Conclusions
We report on the synthesis of uniform-sized mesostructured silica-coated gold nanorods loaded with an organic cyanine dye using physical adsorption or electrostatic adsorption methods. In vivo imaging results using these nanocomposites show for the first time the dual mode imaging capability of a single nanoparticle probe using CT and NIR fluorescence. This novel type of multimodal nanostructure allow the user to more easily obtain multiplexed CT and NIR fluorescence images compared with traditional contrast agents. The application of the dual mode contrast agent in vivo expands the prospects for multiple diagnoses and molecular imaging as well as image guided therapy.
Acknowledgments
This research was supported by the National Basic Research Program of China [No. 2011CB707504, No.2010CB933901, No. 2010CB933903], the Shanghai Commission for Science and Technology, Shanghai, China [No. 09JC1411902], the Research Fund for the Doctoral Program of Higher Education [200802481029], and “111” project from MOE China [No. B08020]. The authors thank Dr. T. FitzGibbon for comments on earlier drafts of the manuscript.
References and links
1. .I. Hizoh and C. Haller, “Radiocontrast-induced renal tubular cell apoptosis: hypertonic versus oxidative stress,” Invest. Radiol. 37(8), 428–434 (2002). [CrossRef] [PubMed]
2. .C. Haller and I. Hizoh, “The cytotoxicity of iodinated radiocontrast agents on renal cells in vitro,” Invest. Radiol. 39(3), 149–154 (2004). [CrossRef] [PubMed]
3. .H. B. Na, I. C. Song, and T. Hyeon, “Inorganic nanoparticles for MRI contrast agents,” Adv. Mater. (Deerfield Beach Fla.) 21(21), 2133–2148 (2009). [CrossRef]
4. .A. H. Lu, E. L. Salabas, and F. Schüth, “Magnetic nanoparticles: synthesis, protection, functionalization, and application,” Angew. Chem. Int. Ed. Engl. 46(8), 1222–1244 (2007). [CrossRef] [PubMed]
5. .J. M. Klostranec and W. C. W. Chan, “Quantum dots in biological and biomedical research: Recent progress and present challenges,” Adv. Mater. (Deerfield Beach Fla.) 18(15), 1953–1964 (2006). [CrossRef]
6. .M. S. Han, A. K. R. Lytton-Jean, B. K. Oh, J. Heo, and C. A. Mirkin, “Colorimetric screening of DNA-binding molecules with gold nanoparticle probes,” Angew. Chem. Int. Ed. Engl. 45(11), 1807–1810 (2006). [CrossRef] [PubMed]
7. .L. L. Ma, M. D. Feldman, J. M. Tam, A. S. Paranjape, K. K. Cheruku, T. A. Larson, J. O. Tam, D. R. Ingram, V. Paramita, J. W. Villard, J. T. Jenkins, T. Wang, G. D. Clarke, R. Asmis, K. Sokolov, B. Chandrasekar, T. E. Milner, and K. P. Johnston, “Small multifunctional nanoclusters (nanoroses) for targeted cellular imaging and therapy,” ACS Nano 3(9), 2686–2696 (2009). [CrossRef] [PubMed]
8. .C. Xu, G. A. Tung, and S. Sun, “Size and concentration effect of gold nanoparticles on X-ray attenuation as measured on computed tomography,” Chem. Mater. 20(13), 4167–4169 (2008). [CrossRef] [PubMed]
9. .P. A. Jackson, W. N. Rahman, C. J. Wong, T. Ackerly, and M. Geso, “Potential dependent superiority of gold nanoparticles in comparison to iodinated contrast agents,” Eur. J. Radiol. 75(1), 104–109 (2010). [CrossRef] [PubMed]
10. .J. F. Hainfeld, D. N. Slatkin, T. M. Focella, and H. M. Smilowitz, “Gold nanoparticles: a new X-ray contrast agent,” Br. J. Radiol. 79(939), 248–253 (2006). [CrossRef] [PubMed]
11. .D. Kim, S. Park, J. H. Lee, Y. Y. Jeong, and S. Jon, “Antibiofouling polymer-coated gold nanoparticles as a contrast agent for in vivo X-ray computed tomography imaging,” J. Am. Chem. Soc. 129(24), 7661–7665 (2007). [CrossRef] [PubMed]
12. .Q. Y. Cai, S. H. Kim, K. S. Choi, S. Y. Kim, S. J. Byun, K. W. Kim, S. H. Park, S. K. Juhng, and K. H. Yoon, “Colloidal gold nanoparticles as a blood-pool contrast agent for X-ray computed tomography in mice,” Invest. Radiol. 42(12), 797–806 (2007). [CrossRef] [PubMed]
13. .R. Guo, H. Wang, C. Peng, M. W. Shen, M. J. Pan, X. Y. Cao, G. X. Zhang, and X. Y. Shi, “X-ray attenuation property of dendrimer-entrapped gold nanoparticles,” J. Phys. Chem. C 114(1), 50–56 (2010). [CrossRef]
14. .V. Kattumuri, K. Katti, S. Bhaskaran, E. J. Boote, S. W. Casteel, G. M. Fent, D. J. Robertson, M. Chandrasekhar, R. Kannan, and K. V. Katti, “Gum arabic as a phytochemical construct for the stabilization of gold nanoparticles: in vivo pharmacokinetics and X-ray-contrast-imaging studies,” Small 3(2), 333–341 (2007). [CrossRef] [PubMed]
15. .H. Wang, L. Zheng, C. Peng, R. Guo, M. Shen, X. Shi, and G. Zhang, “Computed tomography imaging of cancer cells using acetylated dendrimer-entrapped gold nanoparticles,” Biomaterials 32(11), 2979–2988 (2011). [CrossRef] [PubMed]
16. .C. J. Hall, E. Schültke, L. Rigon, K. Ataelmannan, S. Rigley, R. Menk, F. Arfelli, G. Tromba, S. Pearson, S. Wilkinson, A. Round, S. Crittell, R. Griebel, and B. H. J. Juurlink, “Synchrotron-based in vivo tracking of implanted mammalian cells,” Eur. J. Radiol. 68(3Suppl), S156–S159 (2008). [CrossRef] [PubMed]
17. .R. Popovtzer, A. Agrawal, N. A. Kotov, A. Popovtzer, J. Balter, T. E. Carey, and R. Kopelman, “Targeted gold nanoparticles enable molecular CT imaging of cancer,” Nano Lett. 8(12), 4593–4596 (2008). [CrossRef] [PubMed]
18. .W. Eck, A. I. Nicholson, H. Zentgraf, W. Semmler, and S. Bartling, “Anti-CD4-targeted gold nanoparticles induce specific contrast enhancement of peripheral lymph nodes in X-ray computed tomography of live mice,” Nano Lett. 10(7), 2318–2322 (2010). [CrossRef] [PubMed]
19. .D. Kim, Y. Y. Jeong, and S. Jon, “A drug-loaded aptamer-gold nanoparticle bioconjugate for combined CT imaging and therapy of prostate cancer,” ACS Nano 4(7), 3689–3696 (2010). [CrossRef] [PubMed]
20. .J. F. Hainfeld, D. N. Slatkin, and H. M. Smilowitz, “The use of gold nanoparticles to enhance radiotherapy in mice,” Phys. Med. Biol. 49(18), N309–N315 (2004). [CrossRef] [PubMed]
21. .J. F. Hainfeld, F. A. Dilmanian, D. N. Slatkin, and H. M. Smilowitz, “Radiotherapy enhancement with gold nanoparticles,” J. Pharm. Pharmacol. 60(8), 977–985 (2008). [CrossRef] [PubMed]
22. .Z. B. Li, W. Cai, and X. Chen, “Semiconductor quantum dots for in vivo imaging,” J. Nanosci. Nanotechnol. 7(8), 2567–2581 (2007). [CrossRef] [PubMed]
23. .Y. Kong, J. Chen, F. Gao, W. Li, X. Xu, O. Pandoli, H. Yang, J. Ji, and D. Cui, “A multifunctional ribonuclease-A-conjugated CdTe quantum dot cluster nanosystem for synchronous cancer imaging and therapy,” Small 6(21), 2367–2373 (2010). [CrossRef] [PubMed]
24. .C. H. Lee, S. H. Cheng, Y. J. Wang, Y. C. Chen, N. T. Chen, J. Souris, C. T. Chen, C. Y. Mou, C. S. Yang, and L. W. Lo, “Near-infrared mesoporous silica nanoparticles for optical imaging: characterization and in vivo biodistribution,” Adv. Funct. Mater. 19(2), 215–222 (2009). [CrossRef]
25. .W. S. Kuo, C. N. Chang, Y. T. Chang, M. H. Yang, Y. H. Chien, S. J. Chen, and C. S. Yeh, “Gold nanorods in photodynamic therapy, as hyperthermia agents, and in near-infrared optical imaging,” Angew. Chem. Int. Ed. Engl. 49(15), 2711–2715 (2010). [PubMed]
26. .J. S. Souris, C. H. Lee, S. H. Cheng, C. T. Chen, C. S. Yang, J. A. Ho, C. Y. Mou, and L. W. Lo, “Surface charge-mediated rapid hepatobiliary excretion of mesoporous silica nanoparticles,” Biomaterials 31(21), 5564–5574 (2010). [CrossRef] [PubMed]
27. .M. Xiao, J. Nyagilo, V. Arora, P. Kulkarni, D. Xu, X. Sun, and D. P. Davé, “Gold nanotags for combined multi-colored Raman spectroscopy and x-ray computed tomography,” Nanotechnology 21(3), 035101 (2010). [CrossRef] [PubMed]
28. .C. Alric, J. Taleb, G. Le Duc, C. Mandon, C. Billotey, A. Le Meur-Herland, T. Brochard, F. Vocanson, M. Janier, P. Perriat, S. Roux, and O. Tillement, “Gadolinium chelate coated gold nanoparticles as contrast agents for both X-ray computed tomography and magnetic resonance imaging,” J. Am. Chem. Soc. 130(18), 5908–5915 (2008). [CrossRef] [PubMed]
29. .C. Alric, R. Serduc, C. Mandon, J. Taleb, G. Le Duc, A. Le Meur-Herland, C. Billotey, P. Perriat, S. Roux, and O. Tillement, “Gold nanoparticles designed for combining dual modality imaging and radiotherapy,” Gold Bull. 41(2), 90–97 (2008). [CrossRef]
30. .P. Huang, Z. Li, J. Lin, D. Yang, G. Gao, C. Xu, L. Bao, C. Zhang, K. Wang, H. Song, H. Hu, and D. Cui, “Photosensitizer-conjugated magnetic nanoparticles for in vivo simultaneous magnetofluorescent imaging and targeting therapy,” Biomaterials 32(13), 3447–3458 (2011). [CrossRef] [PubMed]
31. .M. M. van Schooneveld, D. P. Cormode, R. Koole, J. T. van Wijngaarden, C. Calcagno, T. Skajaa, J. Hilhorst, D. C. ’t Hart, Z. A. Fayad, W. J. M. Mulder, and A. Meijerink, “A fluorescent, paramagnetic and PEGylated gold/silica nanoparticle for MRI, CT and fluorescence imaging,” Contrast Media Mol. Imaging 5(4), 231–236 (2010). [CrossRef] [PubMed]
32. .X. Huang, I. H. El-Sayed, W. Qian, and M. A. El-Sayed, “Cancer cell imaging and photothermal therapy in the near-infrared region by using gold nanorods,” J. Am. Chem. Soc. 128(6), 2115–2120 (2006). [CrossRef] [PubMed]
33. .L. Tong, Y. Zhao, T. B. Huff, M. N. Hansen, A. Wei, and J. X. Cheng, “Gold nanorods mediate tumor cell death by compromising membrane integrity,” Adv. Mater. (Deerfield Beach Fla.) 19(20), 3136–3141 (2007). [CrossRef] [PubMed]
34. .Z. Li, P. Huang, X. Zhang, J. Lin, S. Yang, B. Liu, F. Gao, P. Xi, Q. Ren, and D. Cui, “RGD-conjugated dendrimer-modified gold nanorods for in vivo tumor targeting and photothermal therapy,” Mol. Pharm. 7(1), 94–104 (2010). [CrossRef] [PubMed]
35. .Y. S. Chen, W. Frey, S. Kim, K. Homan, P. Kruizinga, K. Sokolov, and S. Emelianov, “Enhanced thermal stability of silica-coated gold nanorods for photoacoustic imaging and image-guided therapy,” Opt. Express 18(9), 8867–8878 (2010). [CrossRef] [PubMed]
36. .C. G. Wang, Y. Chen, T. T. Wang, Z. F. Ma, and Z. M. Su, “Monodispersed gold nanorod-embedded silica particles as novel Raman labels for biosensing,” Adv. Funct. Mater. 18(2), 355–361 (2008). [CrossRef]
37. .G. von Maltzahn, A. Centrone, J. Park, R. Ramanathan, M. Sailor, T. Hatton, and S. Bhatia, “SERS-coded Gold nanorods as a multifunctional platform for densely multiplexed near-infrared imaging and photothermal heating,” Adv. Mater. (Deerfield Beach Fla.) 21(31), 3175–3180 (2009). [CrossRef]
38. .C. C. Chen, Y. P. Lin, C. W. Wang, H. C. Tzeng, C. H. Wu, Y. C. Chen, C. P. Chen, L. C. Chen, and Y. C. Wu, “DNA-gold nanorod conjugates for remote control of localized gene expression by near infrared irradiation,” J. Am. Chem. Soc. 128(11), 3709–3715 (2006). [CrossRef] [PubMed]
39. .B. Pan, L. Ao, F. Gao, H. Tian, R. He, and D. Cui, “End-to-end self-assembly and colorimetric characterization of gold nanorods and nanospheres via oligonucleotide hybridization,” Nanotechnology 16(9), 1776–1780 (2005). [CrossRef]
40. .C. Murphy and N. Jana, “Controlling the aspect ratio of inorganic nanorods and nanowires,” Adv. Mater. (Deerfield Beach Fla.) 14(1), 80–82 (2002). [CrossRef]
41. .X. Li, F. J. Kao, C. C. Chuang, and S. He, “Enhancing fluorescence of quantum dots by silica-coated gold nanorods under one- and two-photon excitation,” Opt. Express 18(11), 11335–11346 (2010). [CrossRef] [PubMed]
42. .T. Zhao, H. Wu, S. Q. Yao, Q. H. Xu, and G. Q. Xu, “Nanocomposites containing gold nanorods and porphyrin-doped mesoporous silica with dual capability of two-photon imaging and photosensitization,” Langmuir 26(18), 14937–14942 (2010). [CrossRef] [PubMed]
43. .W. Stöber, A. Fink, and E. Bohn, “Controlled growth of monodisperse silica spheres in the micron size range,” J. Colloid Interface Sci. 26(1), 62–69 (1968). [CrossRef]
44. .B. Nikoobakht, Z. L. Wang, and M. A. El-Sayed, “Self-Assembly Of Gold Nanorods,” J. Phys. Chem. B 104(36), 8635–8640 (2000). [CrossRef]
45. .E. Oh, M. Y. Hong, D. Lee, S. H. Nam, H. C. Yoon, and H. S. Kim, “Inhibition assay of biomolecules based on fluorescence resonance energy transfer (FRET) between quantum dots and gold nanoparticles,” J. Am. Chem. Soc. 127(10), 3270–3271 (2005). [CrossRef] [PubMed]
46. .E. Khon, A. Mereshchenko, A. N. Tarnovsky, K. Acharya, A. Klinkova, N. N. Hewa-Kasakarage, I. Nemitz, and M. Zamkov, “Suppression of the plasmon resonance in Au/CdS colloidal nanocomposites,” Nano Lett. 11(4), 1792–1799 (2011). [CrossRef] [PubMed]
47. .W. Krause, “Delivery of diagnostic agents in computed tomography,” Adv. Drug Deliv. Rev. 37(1-3), 159–173 (1999). [CrossRef] [PubMed]
48. .M. Banna and P. S. Olutola, “Orbital histiocytosis on computed tomography,” J. Comput. Tomogr. 7(2), 167–170 (1983). [CrossRef] [PubMed]
49. .F. Büther, L. Stegger, M. Dawood, F. Range, M. Schäfers, R. Fischbach, T. Wichter, O. Schober, and K. P. Schäfers, “Effective methods to correct contrast agent-induced errors in PET quantification in cardiac PET/CT,” J. Nucl. Med. 48(7), 1060–1068 (2007). [CrossRef] [PubMed]
50. .F. A. Dilmanian, X. Y. Wu, E. C. Parsons, B. Ren, J. Kress, T. M. Button, L. D. Chapman, J. A. Coderre, F. Giron, D. Greenberg, D. J. Krus, Z. Liang, S. Marcovici, M. J. Petersen, C. T. Roque, M. Shleifer, D. N. Slatkin, W. C. Thomlinson, K. Yamamoto, and Z. Zhong, “Single-and dual-energy CT with monochromatic synchrotron x-rays,” Phys. Med. Biol. 42(2), 371–387 (1997). [CrossRef] [PubMed]
51. .C. Xu, G. A. Tung, and S. Sun, “Size and concentration effect of gold nanoparticles on X-ray attenuation as measured on computed tomography,” Chem. Mater. 20(13), 4167–4169 (2008). [CrossRef] [PubMed]
52. .M. Vallet-Regí, F. Balas, and D. Arcos, “Mesoporous materials for drug delivery,” Angew. Chem. Int. Ed. Engl. 46(40), 7548–7558 (2007). [CrossRef] [PubMed]
53. .I. I. Slowing, B. G. Trewyn, and V. S. Y. Lin, “Mesoporous silica nanoparticles for intracellular delivery of membrane-impermeable proteins,” J. Am. Chem. Soc. 129(28), 8845–8849 (2007). [CrossRef] [PubMed]
54. .R. Rajagopalan, P. Uetrecht, J. E. Bugaj, S. A. Achilefu, and R. B. Dorshow, “Stabilization of the optical tracer agent indocyanine green using noncovalent interactions,” Photochem. Photobiol. 71(3), 347–350 (2000). [CrossRef] [PubMed]
