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Langmuir 2007, 23, 9050-9056
Fabrication of Magnetic Core@Shell Fe Oxide@Au Nanoparticles for Interfacial Bioactivity and Bio-separation Hye-Young Park,† Mark J. Schadt, Lingyan Wang, I-Im Stephanie Lim, Peter N. Njoki, Soo Hong Kim, Min-Young Jang, Jin Luo, and Chuan-Jian Zhong* Department of Chemistry, State UniVersity of New York at Binghamton, Binghamton, New York 13902 ReceiVed May 4, 2007. In Final Form: May 30, 2007 The immobilization of proteins on gold-coated magnetic nanoparticles and the subsequent recognition of the targeted proteins provide an effective means for the separation of proteins via application of a magnetic filed. A key challenge is the ability to fabricate such nanoparticles with the desired core-shell nanostructure. In this article, we report findings of the fabrication and characterization of gold-coated iron oxide (Fe2O3 and Fe3O4) core@shell nanoparticles (Fe oxide@Au) toward novel functional biomaterials. A hetero-interparticle coalescence strategy has been demonstrated for fabricating Fe oxide@Au nanoparticles that exhibit controllable sizes ranging from 5 to 100 nm and high monodispersity. Composition and surface analyses have proven that the resulting nanoparticles consist of the Fe2O3 core and the Au shell. The magnetically active Fe oxide core and thiolate-active Au shell were shown to be viable for exploiting the Au surface protein-binding reactivity for bioassay and the Fe oxide core magnetism for magnetic bioseparation. These findings are entirely new and could form the basis for fabricating magnetic nanoparticles as biomaterials with tunable size, magnetism, and surface binding properties.
Introduction The fabrication of magnetic nanoparticles and nanocomposites has attracted both fundamental and practical interest because of potential applications in areas such as ferrofluids, medical imaging, drug targeting and delivery, cancer therapy, separations, and catalysis.1 However, one of the major obstacles is the lack of surface tunability for biocompatible applications. Coating the magnetic particles with a gold shell provides an intriguing class of biomaterials to overcome such an obstacle because the wellestablished surface chemistry and biological reactivity of gold can impart the magnetic particles with the desired chemical or biomedical properties.2 As illustrated in Scheme 1, the immobilization of antibodies on the gold-coated magnetic nanoparticles and the subsequent recognition of specific proteins provide an effective means for the separation of proteins via application of a magnetic field. Similar strategies can also be utilized for DNA separation. A key advantage for putting a gold shell on magnetic particles for the above bioactivity and bioseparation is the ability to exploit the rich surface chemistry of gold while maintaining the magnetic properties. The challenge is the synthesis of core-shell magnetic materials in controllable ways. Several methods for the synthesis of gold-coated magnetic core-shell particles have been reported, including γ-ray radiation, laser ablation, sonochemical reaction, layer-by-layer electrostatic deposition, chemical reduction, and micelle methods.3 We have recently reported a sequential synthesis method to produce gold-coated iron oxide (Fe2O3 and Fe3O4) core@shell nanoparticles (Fe oxide@Au) by the reduction and deposition of gold onto presynthesized iron oxide nano* Corresponding author. E-mail:
[email protected]. † Present address: Institute Pasteur Korea, Seoul, South Korea. (1) (a) Kim, D. K.; Zhang, Y.; Kehr, J.; Klason, T.; Bjelke, B.; Muhammed, M. J. Magn. Magn. Mater. 2001, 225, 256. (b) Niemeyer, C. M. Angew. Chem., Int. Ed. 2001, 40, 4128. (c) Neuberger, T.; Schoepf, B.; Hofmann, H.; Hofmann, M.; Von, Rechenberg, B. J. Magn. Magn. Mater. 2005, 293, 483. (d) Tartaj, P.; Morales, M. P.; Gonzalez-Carreno, T.; Veintemillas-Verdaguer, S.; Serna, C. J. J. Magn. Magn. Mater. 2005, 290, 28. (e) Dobson, J. Drug DeV. Res. 2006, 67, 55. (2) (a) Daniel, M.-C.; Astruc, D. Chem. Rev. 2004, 104, 293. (b) Xia, Y.; Gates, B.; Yin, Y.; Lu, Y. AdV. Mater. 2000, 12, 693. (c) Grubisha, D. S.; Lipert, R. J.; Park, H. Y.; Driskell, J.; Porter, M. D. Anal. Chem. 2003, 75, 5936.
particles.4 This method has proven to produce highly monodisperse Fe oxide@Au nanoparticles. To further refine such core@shell nanoparticles with the desired size range and surface properties, which is often challenging in practical application, we developed another viable approach to the preparation of goldcoated iron oxide (Fe oxide@Au) nanoparticles. This approach applies a thermal processing strategy for the fabrication of Fe oxide@Au magnetic nanoparticles with simple and effective controllability. Scheme 2 illustrates the thermal processing strategy for the fabrication of Fe oxide@Au (Fe2O3@Au or Fe3O4@Au) nanoparticles in which the capping molecules on the Au nanoparticle precursor can be alkanethiolates and those on the Fe oxide (Fe2O3 or Fe3O4) nanoparticle precursor can be oleylamine and/or oleic acid. The basic idea in Scheme 2 explores the viability of thermally activated hetero-interparticle coalescence between gold and magnetic nanoparticles under an encapsulating environment in creating core-shell type nanoparticles. The interior of the coreshell nanoparticles consists of iron oxide cores. Depending on the degree of coalescence and the relative capping agent and precursor concentrations, the formation of both single and multiple magnetic cores inside the shell is possible. This strategy expands the homo-interparticle coalescence demonstrated for the evolution of gold nanoparticles at a mild temperature elevation (140160 °C).5 The basis stems from the decreased melting point, especially for surface melting, for gold at the nanoscale dimension. In contrast, iron oxide nanoparticles do not exhibit any coalescence under such a mild temperature elevation. The competition between growing Au and Fe oxide@Au with single or multiple Fe oxide (3) (a) Kinoshita, T.; Seino, S.; Mizukoshi, Y.; Otome, Y.; Nakagawa, T.; Okitsu, K.; Yamamoto, T. A. J. Magn. Magn. Mater. 2005, 293, 106. (b) Zhang, J.; Post, M.; Veres, T.; Jakubek, Z. J.; Guan, J.; Wang, D.; Normandin, F.; Deslandes, Y.; Simard, B. J. Phys. Chem. B 2006, 110, 7122. (c) Caruntu, D.; Cushing, B. L.; Caruntu, G.; O’Connor, C. J. Chem. Mater. 2005, 17, 3398. (d) Spasova, M.; Salgueirino-Maceira, V.; Schlachter, A.; Hilgendorff, M.; Giersig, M.; Liz-Marzan, L. M.; Farle, M. J. Mater. Chem. 2005, 15, 2095. (e) Stoeva, S. I.; Huo, F.; Lee, J.-S.; Mirkin, C. A. J. Am. Chem. Soc. 2005, 127, 15362. (f) Lyon, J. L.; Fleming, D. A.; Stone, M. B.; Schiffer, P.; Williams, M. E. Nano Lett. 2004, 4, 719. (g) Mandal, M.; Kundu, S.; Ghosh, S. K.; Panigrahi, S.; Sau, T. K.; Yusuf, S. M.; Pal, T. J. Colloid Interface Sci. 2005, 286, 187. (4) Wang, L.; Luo, J.; Fan, Q.; Suzuki, M.; Suzuki, I. S.; Engelhard, M. H.; Lin, Y.; Kim, N.; Wang, J. Q.; Zhong, C. J. J. Phys. Chem. B 2005, 109, 21593.
10.1021/la701305f CCC: $37.00 © 2007 American Chemical Society Published on Web 07/13/2007
Magnetic Core@Shell Fe Oxide@Au Nanoparticles
Langmuir, Vol. 23, No. 17, 2007 9051
Scheme 1. (Top) Idealized Illustration of the Immobilization of the Antibody on the Gold-Coated Magnetic Nanoparticles and Subsequent Recognition of the Proteins and (Bottom) Utilization of This Strategy for the Separation of Proteins via Application of a Magnetic Field
Scheme 2. Idealized Illustration of Hetero-interparticle Coalescence of A and B Nanoparticlesa Leading to Core@Shell Nanoparticles
a For examples, molecularly capped Au (A) and Fe O (B) 2 3 nanoparticles in solution.
cores (pomegranate-like) nanoparticles is determined by a combination of solution temperature, composition, and capping structures. Importantly, the presence of molecularly capped iron oxide nanoparticles in the solution created a unique thermal microenvironment for core-shell coalescence, which serves as an effective strategy for monodisperse Fe oxide@Au nanoparticles of controlled sizes. The Fe oxide@Au magnetic nanoparticles have been demonstrated to be viable for the exploitation of proteinbinding reactivity at the gold surface and magnetic bioseparation via the Fe oxide core. In this article, we report the findings of the investigation of this thermal processing route for the fabrication of core-shell magnetic nanoparticles and interfacial chemical and biological reactivities. The findings have demonstrated the ability to thermally fabricate the gold-coated magnetic core-shell nanoparticles and their utility as biomaterials for a range of potential applications, including biosensing, bioseparation, and targeting. Experimental Section Chemicals. Iron pentacarbonyl (Fe(CO)5), phenyl ether, trimethylamine oxide, decanethiol (DT), tetraoctylammonium bromide (TOA-Br), oleylamine (OAM), oleic acid (OA), trimethylamine oxide dihydrate ((CH3)3NO‚2H2O), bovine serum albumin (BSA), 11(5) (a) Schadt, M. J.; Cheung, W.; Luo, J.; Zhong, C. J. Chem. Mater. 2006, 18, 5147. (b) Maye, M. M.; Zheng, W.; Leibowitz, F. L.; Ly, N. K.; Zhong, C. J. Langmuir 2000, 16, 490. (c) Zhong, C. J.; Zheng, W. X.; F. L. Leibowitz; Eichelberger, H. H. Chem. Commun. 1999, 13, 1211.
mercaptoundecanoic acid (MUA), mercaptobenzoic acid (MBA), dithiobis (succinimidyl propionate) (DSP), and other solvents (hexane, toluene, and ethanol) were obtained from Aldrich and were used as received. Anti-rabbit IgG and dithiobis (succinimidyl propionate) (DSP) were purchased from Pierce. Protein A and gold nanoparticles were obtained from Ted Pella. Synthesis and Preparation. Fe2O3 (γ-Fe2O3) nanoparticles and Au nanoparticles were synthesized by known protocols, whereas the preparation of Fe2O3@Au nanoparticles was based on the new protocol developed in this work. For the synthesis of Fe2O3 nanoparticles, Fe2O3 nanoparticles capped with OA (and/or OAM) were prepared on the basis of the modified procedure reported previously.4 Briefly, 0.74 mL of Fe(CO)5 and 5.3 mL of OA (and/or OAM) in 40 mL of phenyl ether were stirred at 100 °C under an argon purge. The solution was heated to 253 °C and refluxed for 1 h. The solution turned dark brown. After the solution was cooled to room temperature, 1.26 g of (CH3)3NO‚2H2O was added, and the solution was stirred at 130 °C for 2 h. The temperature was increased to 253 °C, and the solution was refluxed for 2 h. The reaction solution was stirred overnight. The resulting nanoparticles were precipitated with ethanol and rinsed multiple times. Finally, particles were dispersed in hexane or toluene. For the synthesis of Au nanoparticles, the standard two-phase method reported by Brust and Schiffrin6 was used. Gold nanoparticles of 2 nm diameter encapsulated with DT monolayer shells (Au2nm-DT) were synthesized. For the preparation of Fe2O3@Au nanoparticles, a modified strategy of the thermally activated processing protocol5 was used. The thermal processing treatment of Au nanoparticles involved molecular desorption, nanocrystal core coalescence, and molecular re-encapsulation processes in the evolution of nanoparticle precursors at elevated temperatures (149 °C). The thermal processing of small monolayer-protected nanoparticles as precursors5 has recently gained increasing interest with respect to nanoparticle processing size and monodispersity.7 In a typical thermal processing treatment, 1.4 mL of Au2 nm-DT and Fe2O3 nanoparticles in toluene with various ratios was placed in a reaction tube. The mixed precursor solution contained Au2 nm-DT, OA- and/or OAM-Fe2O3 nanoparticles, toluene, and TOA-Br. The tube was then placed in a preheated Yamato DX400 gravity convection oven at 149 °C for 1 h. Temperature variation (6) Brust, M.; Walker, M.; Bethell, D.; Schiffrin, D. J.; Whyman, R. J. Chem. Soc., Chem. Commun. 1994, 801-802. (7) (a) Clarke N. Z.; Waters, C.; Johnson, K.A.; Satherley, J.; Schiffrin, D.J.; Langmuir, 2001, 17, 6048-6050. (b) Teranishi, T.; Hasegawa, S.; Shimizu, T.; Miyake, M. AdV. Mater. 2001, 13, 1699-1701. (c) Fan, H.; Gabaldon, J.; Brinker, C. J.; Jiang, Y.B. Chem. Commun. 2006, 2323-2325. (d) Terzi, F.; Seeber, R.; Pigani, L.; Zanardi, C.; Pasquali, L.; Nannarone, S.; Fabrizio, M.; Daolio, S. J. Phys. Chem. B 2005, 109, 19397-19402. (e) Shaffer, A. W.; Worden, J. G.; Huo, Q. Langmuir 2004, 20, 8343-8351. (f) Chen, S. W.; Sommers, J. M. J. Phys. Chem. B 2001, 105, 8816-8820.
9052 Langmuir, Vol. 23, No. 17, 2007 from this set point was limited to (1.5 °C. After the 1 h thermal treatment, the reaction tube was allowed to cool down, and the particles were redispersed in toluene. We note that the above approach can also be used to produce Fe3O4@Au nanoparticles. In this article, we focus on the data for Fe2O3@Au nanoparticles largely because of the availability of systematic data for Fe2O3@Au. The term “Fe oxide” was used to refer to a variety of iron oxides, including Fe2O3 and Fe3O4. Preparation of Nanoparticles Capped with Proteins and SERS Labels. The as-synthesized DT-capped Fe oxide@Au particles were transferred to water by ligand exchange using mercaptoundecanoic acid (MUA) by following a procedure reported by Gittins et al.,8 with a slight modification. The nanoparticles were further modified with DSP for protein coupling by ligand exchange. To a 1 mL solution of MUA-capped Fe oxide@Au particles (6 ng/mL) in borate buffer (pH 8.3), 140 µL of 1 mM DSP was added, and the solution was stirred overnight. The nanoparticles were rinsed by centrifugation, and 20 µL of anti-rabbit IgG (2.4 mg/mL) was added by pipet. After overnight incubation, the particles were centrifuged three times and finally resuspended in 2 mM Tris buffer (pH 7.2) with 1% BSA and 0.1% Tween 80. The same method was also used to coat protein A and BSA onto 80 nm Au nanoparticles. DSP (2.5 µL of a 1 mM solution) was added to 1 mL of Au particles (80 nm, 1 × 1010 particles/mL) and reacted overnight. Borate buffer (40 µL of a 50 mM solution) was added, and either protein A or BSA was added to make the final concentration of protein A or BSA to be ∼25 µg/mL. MBA was used as a Raman label.9 To introduce a spectroscopic label onto the Au nanoparticles modified with either protein A or BSA, an ethanol solution of MBA (10 µg/mL) was added, and the mixture was allowed to react overnight. Finally, the particles were centrifuged three times and finally resuspended in 2 mM Tris buffer (pH 7.2) with 1% BSA and 0.1% Tween 80. The resulting protein-capped nanoparticles were stored at 4 °C. Measurements and Instrumentation. The study of the binding between Au nanoparticles labeled with protein A (or BSA) and MBA (A) and Fe oxide@Au nanoparticles labeled with anti-rabbit IgG (B) was carried out by mixing A and B. Solution A (250 µL) was first diluted in 1750 µL of Tris buffer before mixing with ∼10 µL of solution B. UV-vis spectra were collected immediately after gentle mixing of the solution. Spectroscopic measurements were performed after using a magnet to collect the reaction product. For spectroscopic labeling, 0.8 mL of an ethanol solution of MBA was added to 0.5 mL of Fe oxide@Au particles (30 mg/mL in toluene), and the mixture was shaken overnight. The particles were cleaned three times with toluene and ethanol and dispersed in a 2 mM borate buffer. The particles were then drop cast onto gold on a mica surface. Surface-Enhanced Raman Scattering (SERS). Raman spectra were recorded using the Advantage 200A Raman instrument (DeltaNu). The instrument collects data over 200 to 3400 cm-1. The laser power was 5 mW, and the wavelength of the laser was 632.8 nm. In the experiment, we collected the spectrum in the range from 200 to 1800 cm-1. Transmission Electron Microscopy (TEM). TEM micrographs of the particles were obtained using a Hitachi H-7000 electron microscope operated at 100 kV. The particles dispersed in hexane were drop cast onto a carbon-film-coated copper grid, followed by evaporation at room temperature. Ultraviolet-Visible Spectroscopy (UV-Vis). UV-vis spectra were acquired with an HP8453 spectrophotometer. The spectra were collected over the range of 200-1100 nm. Direct Current Plasma Atomic Emission Spectroscopy (DCPAES). The composition of synthesized particles and thin films was analyzed using DCP-AES. Measurements were made on emission peaks at 267.59 and 259.94 nm for Au and Fe, respectively. The nanoparticle samples were dissolved in concentrated aqua regia and then diluted to concentrations in the range of 1 to 50 ppm for analysis. Calibration curves were constructed from standards with concentra(8) Gittins, D. I.; Caruso, F. ChemPhysChem 2002, 3, 110-113. (9) Ni, J.; Lipert, R. J.; Dawson, G. B.; Porter, M. D. Anal. Chem. 1999, 71, 4903.
Park et al.
Figure 1. TEM micrographs of gold nanoparticles produced by the thermal processing (149 °C) of Au2 nm-DT nanoparticles. (a) Precursor nanoparticles (2.0 ( 0.4 nm) and (b) product nanoparticles (6.4 ( 0.4 nm). tions from 0 to 50 ppm in the same acid matrix as for the unknowns. Detection limits, based on three standard deviations of the background intensity, are 0.008 and 0.005 ppm for Au and Fe, respectively. Standards and unknowns were analyzed 10 times each for 3 s counts. Instrument reproducibility, for concentrations greater than 100 times the detection limit, results in
