Monitoring the Covalent Binding of Quantum Dots to Functionalized

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J. Phys. Chem. C 2007, 111, 10313-10319

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Monitoring the Covalent Binding of Quantum Dots to Functionalized Gold Surfaces by Surface Plasmon Resonance Spectroscopy Petra J Cameron,*,†,‡ Xinhua Zhong,§ and Wolfgang Knoll† Max Plank Institute for Polymer Research, Ackermannweg 10, 55128 Mainz, Germany, and Department of Chemistry, East China UniVersity of Science and Technology, 200237 Shanghai, China ReceiVed: January 19, 2007; In Final Form: May 4, 2007

Mercapto-propionic acid-capped cadmium selenide nanoparticles were covalently bound to the terminal amino groups of 11-amino-1-undecanethiol self-assembled monolayers (SAM) on the surface of gold electrodes. The nanoparticle binding was followed by surface plasmon resonance spectroscopy (SPR), and the minimum angle shift was related to surface coverage using atomic force microscopy (AFM) and scanning electron microscopy (SEM). An S-shaped binding isotherm was obtained; in all cases binding saturated at submonolayer coverage. Submonolayer formation was attributed to an exponential fall in the association affinity constant, ka, as a function of surface coverage. Effective medium theory was used to extract dielectric constants (λ ) 632.8 nm) of  ) 3.4 for the MPA-capped CdSe nanoparticles and  ) 1.76 for the surrounding aqueous media.

Introduction Inorganic nanoparticles, also called quantum dots or q-dots, are versatile particles with applications in areas as disparate as biosensing, photovoltaics, “smart” composite materials and nanoelectronics.1-3 They are made from semiconducting materials, typically metal (e.g., Cd, Zn, Pb) chalcogenides. The nanoparticles can be tailored to have diameters ranging from one nanometer to tens of nanometers and their outer ‘shell’ of capping ligands has been chemically modified to carry a host of functional groups, e.g., for attachment to biological molecules.4 The properties of nanoparticles are often unique, resembling neither those of the bulk material nor the individual constituents.4 Changing the size of semiconductor nanoparticles, e.g., in the case of CdSe q-dots, leads to large changes in the band gap energy and allows the fluorescence emission to be tuned across the entire visible spectrum. While lithographic patterning methods struggle to create structures much below 100 nm,5 it is relatively easy to use a “bottom-up” approach to assemble nanometer-sized components into larger patterns and arrays. Quantum dots covalently bound or adsorbed on gold and indium tin oxide electrodes have been investigated in order to improve understanding of the properties of the q-dots, in particular, the nature of charge transfer between nanoparticle and electrode.6-11 One interesting discovery is that the optical properties of the nanoparticles depend both on the nature of the electrode material and on the electron occupation of the individual particles.12-15 Quantum dot arrays are beginning to be considered as platforms for biosensing.2,16-18 Finally, there is interest in “quantum dot solids” (multilayer assemblies of q-dots) for applications as novel lasing media and potential materials for use in light emitting diodes.19,20 * Corresponding author. E-mails: [email protected], [email protected]. † Max Plank Institute for Polymer Research. ‡ Present address: Department of Chemistry, University of Bath, Bath BA2 7AY, United Kingdom. § East China University of Science and Technology.

Various authors have attached nanoparticles (CdSe, CdS, PbSe) to gold and platinum electrodes using di-thiol linkers.21,7,11 In a series of elegant experiments Bakkers et al.11 attached bis-(2-ethylhexyl)sulfosuccinate capped CdSe nanoparticles to gold electrodes using di-thiol and di-thiane linkers. The thiol functionalized gold electrodes were dipped in an aqueous solution of CdSe for one week after which the surface was uniformly covered with CdSe q-dots. By using a range of different spacers, the authors were able to measure the distance-dependent electron tunneling from the nanoparticle to the electrode. More recently, EDC coupling was used to tether amine functionalized CdSe nanoparticles to carboxylic acid functionalized gold nanoparticles and carboxylic acid functionalized carbon nanotubes at electrode surfaces.8,18 The gold nanoparticle-CdSe assemblies were used to detect acetylcholine esterase; the carbon nanotubes were used to enhance charge separation by accepting photo generated electrons from photoexcited quantum dots. In the research described here, carboxylic acid terminated nanoparticles were covalently coupled to amine terminated selfassembled monolayers at gold surfaces. EDC coupling has become a standard method for coupling q-dots to proteins or other biomolecules to act as fluorescent markers.22 EDC reacts with carboxylic acid groups to form a reactive ester, when the ester is introduced to a nucleophilic group such as the primary amines used here, an amide bond forms releasing a urea derivative as a stable leaving group. The system was chosen as the amide bond is readily formed in an aqueous environment and under mild coupling conditions; in addition the coupling reaction is reasonably fastsit was not necessary to incubate the gold samples for long periods in the CdSe solution. Nanoparticle binding was followed in real time by SPR, which allowed kinetic information about the coupling reaction to be extracted. It was possible to control the degree of nanoparticle binding and as a result to obtain information about the dielectric constant as a function of the number of nanoparticles at the surface.

10.1021/jp0704766 CCC: $37.00 © 2007 American Chemical Society Published on Web 06/23/2007

10314 J. Phys. Chem. C, Vol. 111, No. 28, 2007 Experimental Section Materials. Trioctylphosphine (TOP, 90%), oleylamine (97%), oleic acid (90%), 1-octadecene (ODE, 90%), CdO (99.999%), Se powder (99.999%), and 3-mercaptopropionic acid (MPA, 99%) were purchased from Aldrich and used as received. 11Amino-1-undecanethiol (Dojindo), 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC, Sigma Aldrich), N-2-hydroxyehtylpiperazine-N′-2-ethanesulfonic acid (HEPES, Fluka Biochemica, 99.5%), sodium sulfite, sodium hydroxide, and ethanolamine hydrochloride (Acros organics, >99%) were all used as received. Water was purified using a Millipore system and had a minimum resistivity of 18.2MΩ. Synthesis of MPA-Capped CdSe nanoparticles. Oil-soluble CdSe nanoparticles were prepared according to a literature method.23 Typically, 5.0 mL of oleylamine and 0.15 mL of Se stock solution (2.1 M in TOP) were loaded in a 50 mL threeneck round-bottom flask, and the mixture was heated to 300 °C in a flow of argon. 1.0 mL of Cd stock solution (0.3 M, obtained by dissolving CdO in 6-fold of oleic acid and ODE at elevated temperature) was injected quickly into the reaction flask. The temperature was then set at 280 °C for the subsequent growth and annealing of nanocrystals. After completion of particle growth, the reaction mixture was allowed to cool to ∼60 °C, and 10 mL of methanol was added. The obtained CdSe nanocrystals were precipitated by adding methanol into the toluene solution and further isolated and purified by repeated centrifugation and decantation. MPAcapped water-soluble QDs were obtained by a ligand replacement reaction.24 Scanning Electron Microscopy and Transmission Electron Microscopy. SEM was carried out on a 1530 Gemini electron microscope (LEO) and TEM was done with a Tecnai F20 electron microscope. TEM showed particle diameters of ∼5 nm. Atomic Force Microscopy. AFM images were taken by a dimension D3100 from Veeco Instruments. All scans were measured in tapping mode in air. The tips were Omlac 160 TSW2 silicon tips with a diameter in the region of 10 nm. Given that the diameter of the tip was larger than that of the individual nanoparticles, it was not possible to resolve individual nanoparticles in the x-y image. Cross section and depth analysis suggested an average feature height of 4.8 ( 0.5 nm, consistent with a monolayer of nanoparticles forming at the surface. UV-Visible Spectroscopy and Fluorescence Spectroscopy. UV-vis spectroscopy was carried out on a Perkin-Elmer Lambda 9 UV/vis/NIR spectrophotometer and fluorescence spectroscopy on a J&M TIDAS fluorescence spectrometer with illumination from a 100 W xenon bulb. The MPA-capped CdSe quantum dots displayed an absorption onset at 630 nm, although the absorption remained very low until ∼625 nm. It was therefore assumed that the 632.8 nm HeNe laser did not excite band gap transitions in the quantum dots. The nanoparticles showed weak fluorescence centered on 610 nm when excited at 300 nm. Electrochemistry. Electrochemical studies were carried out using an Autolab PGStat 30. All electrochemistry and impedance spectroscopy was carried out in a 0.1M solution of Na2SO3 adjusted to pH 12 with sodium hydroxide. The working electrode was a gold-coated LaSFN9 glass slide (Schott) and the active area was 0.5 cm.2 The counter electrode was a coil of platinum wire and the reference was a silver/silver chloride electrode (Dri-Ref). Short circuit photocurrents were measured under illumination from a high-intensity Luxeon V Lumiled light

Cameron et al. emitting diode (λ ) 470 nm). The beam was focused onto the sample with a collimating lens, and a second lens was used to obtain uniform illumination over the cell area. The incident photon flux (measured using a Newport 1930c power meter) was in the region of 1016 cm-2 s-1. The photocurrent was measured with the Autolab PGStat 30. Surface Plasmon Resonance Spectroscopy. Surface plasmon resonance spectroscopy (SPR) was measured in the Kretschmann attenuated total internal reflection configuration on a home-built setup. Laser (Uniphase, HeNe λ ) 632.8 nm) light was passed through a chopper and two polarizers before being incident on one face of a LaSFN9 prism (Schott Glass). The chopper modulated the light at 431 Hz and provided a reference signal for the lock-in amplifier. The first polarizer was used to adjust the intensity of the incident light and the second one to ensure that only p-polarized light reached the sample. The film of interest was formed on top of a ∼45 nm gold film deposited on a LaSFN9 glass slide (Schott Glass) which was separated from the back of the LaSFN9 prism by a thin layer of index matching fluid (Cargille Laboratories Inc., n ) 1.700 ( 0.0002). The sample and the prism were mounted on a computer-operated goniometer, which was used to control the precise angle of incidence of the light. The reflected light beam was focused through a collecting lens onto a silicon photodiode. A computer program, designed in house, was used to measure the magnitude of reflected light reaching the photodiode as a function of the incident angle controlled by the goniometer. The SPR curves were fit using an iterative fitting program based on the Fresnel equations. Self-Assembled Monolayers on Evaporated Gold Surfaces. Two types of gold substrate were used in experiments. The first set of substrates consisted of gold films thermally evaporated onto clean LaSFN9 glass. In all cases, a 2 nm layer of chromium was evaporated just prior to gold deposition to improve adhesion of the gold layer. The evaporation was carried out using an Edwards Auto 306 evaporator; the gold films had an average thickness of 45-50 nm. These substrates were used for surface plasmon resonance and electrochemical studies. In order to measure AFM and SEM images of the quantum dots on the electrode surface a second set of “template stripped gold” substrates were prepared. Silicon wafers were cleaned for 1 h in a 5:1:1 solution of water:ammonia hydrogen peroxide at 70 °C. A 45-50 nm layer of gold was then evaporated directly onto the silicon wafer. Epoxy glue was degassed for 1 h on a vacuum line and used to stick the gold side of the silicon wafer to a clean glass microscope slide. The glue was allowed to dry at 150 °C for 1 h, and the samples were left overnight to cool down. Excess glue was removed from the edges with a sharp knife and the silicon wafer was “stripped” away in one piece leaving a flat gold film glued onto the microscope slide. The surface roughness of the gold was measured by AFM to be of the order of a few nanometers, flat enough to allow imaging of the 5 nm nanoparticles. The template stripped gold slides were also used in a study to relate the angle shift measured by SPR directly to the surface coverage measured by AFM/ SEM. Self-assembled monolayers of 11-Amino-1- undecanethiol were formed by leaving the gold surface in contact with a 1 mM solution of the thiol in ethanol overnight. The SAM coverage was estimated to be about 91% from impedance spectroscopy and the SAM thickness was calculated to be 1.3 nm.25,26 The SAM coated slides were rinsed in ethanol and dried in a stream of air before being fixed in place as one wall of a

Covalent Binding of Quantum Dots

Figure 1. Surface plasmon resonance curves before (circles) and after (triangles) binding ∼0.29 of a monolayer of CdSe nanoparticles. The solid lines show fits based on the Fresnel equations. The fitting parameters are outlined in Table 1.

Teflon flow cell. All binding experiments and electrochemical measurements were thereafter done in situ in a combined electrochemical/SPR flow cell. Covalent Coupling of Nanoparticles to Amine Functionalized Surfaces. The required concentration of CdSe nanoparticles was added to a 2 mM solution of 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) in HEPES buffer at pH 7.5 to create nanoparticles with reactive EDC-ester groups at the surface. After 5 min, the solution was allowed to flow through the Teflon flow cell and across the 11-amino-1-undecanethiol SAM at a rate of 1.8 mL/minute. The EDC/CdSe solution was recycled around the closed flow system until no more binding was observed. 0.1M ethanolamine was introduced to the cell to remove any unreacted ester. Finally, the cell was rinsed through with more HEPES buffer. Results and Discussion Formation of CdSe Layers on Gold. The covalent coupling of CdSe quantum dots to amine functionalized gold electrodes was followed in real time by surface plasmon resonance spectroscopy (SPR). The surface coverage was extrapolated from changes in the position of the plasmon minimum angle observed as the ∼5 nm diameter CdSe nanoparticles replaced the buffer solution close to the gold surface. Typical curves before and after binding a partial monolayer of nanoparticles can be seen in Figure 1. In order to obtain a more accurate idea of the relationship between the minimum angle shift measured by SPR and the resulting fractional CdSe surface coverage, a series of six samples were prepared on template stripped gold. These substrates had an average surface roughness 18 h). Both AFM and SEM showed that the fractional coverage of CdSe nanoparticles on template stripped gold measured by surface imaging techniques was linearly related to the minimum angle shift measured by SPR. A linear calibration plot was constructed and used to predict fractional surface coverage in further experiments with gold coated LaSFN9 slides, assuming that a similar relationship between surface coverage and minimum angle shift occurred on both the template-stripped gold (roughness 1-2 nm) and gold-coated LaSFN9 (roughness 15-20 nm). Dielectric Constant of CdSe Nanoparticles. An iterative fitting program based on the Fresnel equations was used to fit the angular SPR scans before and after nanoparticle binding (λ ) 632.8 nm). The angle scans for the gold slides covered with 11-amino-1-undecanethiol SAMs were fit using a five-layer model (glass slide |chrome layer|gold layer |11-amino-1undecanethiol film|solvent layer). It was possible to achieve a good fit (solid line through open circles in Figure 1) based on literature values27-29 of the optical properties for each layer and the thickness of the chrome and gold layers measured during deposition. The values of the dielectric constants used for fitting are outlined in Table 1. The second scan, taken after CdSe binding, was fit by adding an extra layer to the model (CdSe layer was inserted between layers 4 and 6 as outlined in Table 1). The dielectric properties of the CdSe film were input and varied, while the optical constants and thickness of all other layers were held constant. The dielectric properties of the CdSe layer are given by the complex function  ) ′ + i′′, where ′ is the real part of the dielectric and ′′ the imaginary part. The extinction coefficient of the CdSe nanoparticles is negligible at 632.8 nm, so the dielectric function was taken to be real. When ′′ is neglected, the real dielectric component equals the square root of the refractive index of the CdSe film, n. The CdSe layer was composed of organic molecule capped CdSe particles and solvent filled voids. The effective dielectric constant of the layer, eff, was therefore some combination of the dielectric constants of the nanoparticles (np) and the interpenetrating medium (solvent, sol). The thickness of the CdSe layer was set to 4.8 nm and a good fit was obtained by varying only the real part of the dielectric (solid line through open triangles in Figure 1). Values of eff were obtained for a range of samples with different fractional CdSe coverage. The Bruggeman expression, also know as the effective medium expression (EMA), describes the average or effective dielectric constant of a heterogeneous mixture (on the macro scale) of two different dielectric media (A and B).30 It assumes that inclusions of one dielectric material, A, are embedded in an averaged dielectric medium (A + B). It is particularly applicable when the volume fractions of A and B are similar. Hutter et al.21 used the EMA to describe the dielectric properties of CdS nanoparticles attached to self-assembled monolayers on gold. The authors measured the eff of CdS layers in air; np could not be extracted as not enough information was known about the fractional surface coverage. The EMA is given by eq 1a, where eff is the effective dielectric constant for the mixed nano-

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Figure 2. Atomic force microscope images showing height (left-hand images) and phase (corresponding right-hand images) of (A) templatestripped gold covered in a self-assembled monolayer of 11-amino-1-undecanethiol, (B) 19% coverage of CdSe nanoparticles, and (C) 63% coverage of CdSe nanoparticles. At low coverage the nanoparticles appear to group into island on the surface. Although some 3-D aggregation does occur, especially when depositing higher concentrations, AFM analysis gave an average feature height of 4.8 ( 0.5 nm, consistent with a monolayer at the surface.

particle/solvent layer, np is the dielectric constant for the nanoparticles, fnp is the volume fraction of nanoparticles,

sol is the dielectric constant for the solvent, and fsol is the volume fraction of solvent:

Covalent Binding of Quantum Dots

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sol - eff (np - eff) + fsol fnp )0 sol + 2eff (np + 2eff)

(1a)

Assuming that fsol + fnp)1, the EMA can be rearranged to:

fnp

)

(fnp - 1)

(sol - eff)(np - 2eff) (np - eff)(sol - 2eff)

(1b)

In the case of the CdSe layers described here, it was possible to use effective medium theory to extract both np and sol. Figure 3 shows a plot of eff as a function of fnp/(fnp - 1) for a series of values of fnp (ranging from 0.01 to 0.69). The data was fit to eq 1b (solid line in Figure 3), allowing both np and sol to vary. The best fit was obtained for sol ) 1.76 and np ) 3.4; it is encouraging that sol (for dilute aqueous buffer solution) was found to be very close to the literature value of the dielectric constant of water at 632.8 nm (1.77).31 np was found to be lower than the dielectric constant for bulk crystalline CdSe, which is in the region of 5.4 at 638.2 nm.32 Both the tight binding approximation and the effective mass approximation are frequently used to calculate the optical properties of semiconductor nanocrystals.33-36 There is still controversy over what value of the dielectric constant should be input to obtain results that best fit the experimental data; with some models using values for the static dielectric constant, (0) (low-frequency limit of ), and others values for the optical dielectric constant, (∞) (high-frequency limit of ). In addition, a lack of experimental data for the dielectric constants of nanocrystals leads to values for the bulk crystalline material being used to describe the nanocrystalline material. Semiempirical and ab initio calculations of the dielectric constant of silicon quantum dots have suggested that the nanocrystals should display lower effective dielectric constants than the bulk material due to the influence of the particle surface on the dielectric response.37-39 One model, developed by Haken, uses a combination of (0) and (∞) to calculate a size dependent dielectric constant, SD, for semiconductor nanoparticles.34 The model also predicts a decrease in SD with decreasing particle size. In the work described here it was possible to extract an experimental value of the dielectric constant of the nanoparticles themselves, albeit at a single wavelength (λ ) 632.8 nm). The calculated value is considerably lower than the literature values for bulk crystalline CdSe, but it does encompass both the nanocrystal and its accompanying ligand shell. A rough estimate of the dielectric constant of the nanocrystal in the absence of ligands can be made, by assuming a 0.4 nm organic shell with a dielectric constant of 2.25 surrounding the crystalline center. If the calculated dielectric constant of 3.4 is assumed to be a linear combination of nanocrystal and shell, the dielectric constant of the crystalline center is approximately 4.2. While this is a rough estimate, it shows that the dielectric constant is still considerably lower than literature values for bulk CdSe at this wavelength. Kinetics of Nanoparticle Binding. Titration experiments were carried out to investigate the concentration dependence TABLE 1: Film Thickness and Dielectric Constants Used to Fit SPR Angle Scans 1 2 3 4 5 6

layer

thickness

′

′′

LASFN9 prism chrome gold C11-spacer thiol CdSe layer aqueous buffer

1 nm 46 nm 1.3 nm 4.8 nm

3.386 -4 -12.12 2.25 variable 1.77

0 18 1.55 0 0 0

Figure 3. Effective dielectric constant as a function of nanoparticle coverage (open circles); the solid line is the fit obtained with eq 1b when both np and sol were allowed to vary freely.

Figure 4. Changes in the reflectivity (triangles) measured at a single angle of incidence when titrating in CdSe nanoparticles at concentrations between 2 × 10-8 and 10 × 10-8 M (in all cases the CdSe was mixed with 2 mM EDC in HEPES buffer at pH 7.5). The line represents the fit to simple second-order association at the surface.

of the binding kinetics of the CdSe nanoparticles. In the titration experiment CdSe solutions of increasing concentration were added to the flow cell and, at each concentration, binding was measured until equilibrium was reached. Surface plasmon resonance spectroscopy was used to monitor the binding in real time by measuring changes in reflectivity at a fixed angle of incidence (Figure 4). The degree of binding was found to be reproducible providing that all the gold substrates had been deposited simultaneously and that the CdSe nanoparticles came from the same synthetic batch. Figure 5 shows a representative S-shaped binding isotherm, half-saturation coverage was observed at a solution concentration of 7.9 × 10-8 M. The solution concentration of the nanoparticles was estimated from the absorbance at the first exciton absorption peak.40 The binding data shown in Figure 4 were modeled and fit using the expression for simple bimolecular association at a surface (eq 2). Rt is the response at time t, in this case the reflectivity change measured by SPR. Reqm is the response at equilibrium for a given concentration of CdSe nanoparticles in the binding solution:

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Figure 5. S-shaped binding isotherm for the EDC coupling of quantum dots to the terminal amine groups of an 11-amino-1-undecanethiol monolayer. The line is a guide for the eye. The crosses represent points taken from the titration experiment in Figure 4, for each concentration of CdSe added the total change in reflectivity after equilibrium is reached is related to the fraction of CdSe attached to the surface. The crossed open circles represent three independent binding experiments where a single concentration of nanoparticles was bound. The fractional CdSe surface coverage was reproducible providing that the gold slides were prepared at the same time and the nanoparticles come from the same synthetic batch. Two separate titration experiments for two gold slides evaporated in the same batch gave very similar results.

Rt ) Reqm(1 - e-kont)

(2)

kon ) ka[nanoparticles] + kd

(3)

The rate constant for dissociation, kd, was set to zero as the binding was irreversible. This simple model gave a good fit to the data (solid lines in Figure 4) and allowed values for the association constant, ka, to be calculated. The variation in ka with concentration of CdSe in the binding solution is shown in Figure 6. As the coverage of the CdSe at the surface increased, the association constant fell off exponentially. The result contrasts with the Langmuir model of surface adsorption where the assumption is made that ka is constant with concentration, that is to say there is negligible interaction between bound molecules and incoming molecules.41 An exponential fall in ka with concentration suggests that not all the binding sites were equally attractive to incoming particles. However, if the exponential decay in the association constant were due to simple repulsion of single incoming particles by bound particles, then the bound particles would not group together on the surface as observed by AFM. This observation, in conjunction with the S-shaped binding isotherm, suggests that a more complex mechanism is at work; with the possibility that two or more competing mechanisms are responsible for the binding kinetics. One possible interpretation of the data is that after the reaction of the nanoparticles with EDC to replace some of the carboxylate surface groups with esters, the surface charge of the nanoparticles is decreased and they can associate in solution (indeed several hours after adding EDC to a nanoparticle solution, precipitation of aggregated nanoparticles out of the solution can be observed). The small aggregates of two-three nanoparticles then bind on the surface. When binding is stopped at low coverages (e.g., 19% coverage observed in Figure 2) the nanoparticles appear to be grouped into islands on the surface. At higher coverages however the islands appear to spread out and the spaces between them are filled to some extent. It is

Figure 6. The variation in the association constant, ka, with concentration (open circles). The ka values are obtained from the fits shown in Figure 4. The data points are fit to a single-exponential decay (line).

possible that the steep increase in surface coverage around the half saturation concentration of 7.9 × 10-8 M that is seen in the binding isotherm (Figure 5) is due to a phase transition; possibly a rearrangement occurs to allow more ordered packing of the nanoparticles at the surface, from islands to “strings’. This theory is supported by AFM (see also Figure 2) data which showed small isolated groups of particles on the surface when the binding was terminated at low coverages and smaller more spread out strings/groups of particles when the binding was allowed to go to saturation. It is also possible that the association rate constant falls with coverage as once the clusters of surface bound nanoparticles reach a certain size, the net charge is sufficiently large to repel the incoming aggregates. The fall in ka with coverage also explains why monolayers were observed under these coupling conditions, it was kinetically unfavorable to form multilayers or to continue to pack more nanoparticles onto the surface beyond a certain point. The random sequential adsorption (RSA) model is often used to describe the absorption of proteins and colloids onto surfaces.42 The adsorbing molecules are charged, once bound they create an exclusion zone preventing other particles from binding close by. Experimentally a ‘jamming limit’, or maximum coverage, is observed for monodisperse particles at a surface coverage in the region of 0.57. It is possible that a similar mechanism is at work here, although with groups of nanoparticles rather than single nanoparticles repelling each other. As has been mentioned previously, the surface coverage was observed to vary between gold substrates. On the evaporated gold substrates, the average final saturation surface coverage was 35%, but the standard deviation was high ((15% over 11 experiments). On the template stripped gold substrates the saturation coverage was higher, on several occasions coverage greater than 60% was achieved. It is not yet understood why such a large variation in coverage was measured, but it is feasible that it is related to the surface structure of the gold and the packing of the nanoparticles on the surface. The surface roughness of the gold on the LaSFN9 substrates was of the order of 15-20 nm, much greater than the size of the individual nanoparticles. It is possible that the number of binding sites was dictated by the structure of gold crystallites at the surface. Given the rapid fall in the association constant with coverage, the position of the initial binding sites would have large effects on the final saturation coverage of the film.

Covalent Binding of Quantum Dots

J. Phys. Chem. C, Vol. 111, No. 28, 2007 10319 Acknowledgment. PJC thanks Dr. Jason Riley and Dr. Toby Jenkins for useful discussions, Andreas Scheller for technical support and Gunnar Glassar and Ingo Lieberwirth for SEM and TEM support. PJC acknowledges the Alexander von Humboldt Stiftung for funding. References and Notes

Figure 7. Chopped illumination from a blue LED superimposed on a linear sweep voltammogram at a scan rate of 5 mV/s. The electrode area was 0.5 cm.2

Photocurrent Measurements of CdSe Assemblies under Illumination. A blue (λ ) 470 nm) LED was focused on the CdSe film from the solution side. Photocurrents were measured in three-electrode mode under chopped illumination. The electrolyte was 0.1M Na2SO3 at pH 12. Under illumination charge separation could be measured as a photocurrent in the external circuit. Reference 11 gives a detailed account of electron transfer from quantum-dots to gold electrodes; briefly, light of energy greater than the band gap in the nanoparticle/ q-dot creates an excited state, which can decay via intraparticle recombination of the electron and hole or via transfer of the electron to the gold electrode. If a redox donor (also called a “hole scavenger”) is present in the solution the hole can be rapidly filled and the excited electron can be measured as a photocurrent in the external circuit. Figure 7 shows a typical current-voltage plot at a surface coverage of 25% (∼7 × 1011 nanoparticles), the photocurrent upon illumination can clearly be seen above the background current. The incident light (470 nm) had a photon flux of 1016 photons cm-2 s-1, the resulting short circuit photocurrent was 8.8 nA cm-2. As the molar absorption coefficient of the CdSe nanoparticles in solution is 8 × 106 Lmol-1cm-1 at 470 nm (from UV-vis absorption measurements); this translates to a quantum yield of 0.13%, assuming that the light passes through the nanoparticle layer twice due to the reflecting gold substrate and that the absorption coefficient is the same on the surface as in solution. Conclusions EDC coupling has been shown to be an effective way to couple semiconductor nanoparticles to amine terminated selfassembled monolayers at gold electrodes. SPR was used to investigate the kinetics of nanoparticle binding. The binding kinetics are complex, possibly due to the pre-aggregation of small clusters of nanoparticles in solution that then bind and spread out on the surface. The formation of submonolayers at the surface can be explained by an exponential fall in the association binding constant, ka, with coverage. The effective medium approximation was used to extract a value of the dielectric constant, np, of the ∼5 nm diameter ligand capped nanoparticles at 632.8 nm of 3.4.

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