Precision Patterning with Luminescent Nanocrystal-Functionalized

Aug 5, 2010 - TEM images of the nanocrystals before and after the phase transfer and of the PS beads at every stage of the assembly were recorded...
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Precision Patterning with Luminescent Nanocrystal-Functionalized Beads )

Elisabetta Fanizza,*,†,‡ Laurent Malaquin,§,z Tobias Kraus,§,# Heiko Wolf,§ Marinella Striccoli,‡ Norberto Micali, Antonietta Taurino,^ Angela Agostiano,†,‡ and M. Lucia Curri‡ Department of Chemistry, University of Bari, Via Orabona 4I, Bari 70126, Italy, ‡IPCF-CNR Bari Division, Via Orabona 4I, Bari 70126, Italy, §IBM Research - Zurich, S€ aumerstrasse 4, 8803 R€ uschlikon, Switzerland, CNR- IPCF Messina Division, Contrada Papardo, Salita Sperone, Faro Superiore, 98158 Messina, Italy, and ^ CNR-IMM Lecce Division, Campus Universitario, Strada Prov. Lecce-Monteroni km 1,200, 73100 Lecce, Italy. z Present address: Institut Curie, UMR 168 11 rue Pierre et Marie Curie, 75248 Paris Cedex 5, France. # Present address: Leibniz Institute for New Materials (INM), Campus D2 2, 66123 Saarbr€ ucken, Germany.


Received June 8, 2010. Revised Manuscript Received July 7, 2010 A reliable strategy is presented to combine the preparation of functional building blocks based on polymer beads decorated with luminescent nanocrystals (NCs) and their precise positioning onto suitable patterns by capillary assembly technique. In particular, a layer-by-layer (LbL) polyelectrolyte (PE) deposition procedure has been implemented to provide uniform NC coverage on PS beads, thus conveying the optical properties of luminescent nanocrystals to highly processable PS beads. The latter have then been integrated into patterned stamps by means of template-driven capillary assembly. Their selective positioning has been directed by means of pattern geometry. The use of luminescent (CdSe)ZnS NCs offers a direct optical probe to evaluate the efficiency of the positioning procedure on the substrate, enabling the extension of the method to a wide range of materials, i.e., NCs with different compositions and specific geometry-dependent properties. Moreover, the precise control over the pattern geometry and the micrometer accuracy in positioning achieved by capillary assembly make such functional patterned structures excellent candidates for integration into devices exploiting specific size-dependent NC properties.

Introduction The control over single-particle positioning has recently attracted growing interest, as it represents a powerful opportunity for the fabrication of 2D and 3D architectures and the integration of nanoparticles into micro-nanosystems and devices. The characteristics of such architectures depend on the final arrangement of the nanometer-sized building blocks on the solid surface. The development of procedures to tailor and/or tune the optical, magnetic, and electrical properties of NPs1 and recent advances in micropatterning2 and self-assembly techniques3 provide unique combinations for the fabrication of innovative optoelectronic devices, photonic band gap materials, sensors, and biochips. Several approaches have been reported to selectively drive NP positioning and organization on selected substrates, such as *Corresponding author. E-mail: [email protected]. (1) (a) Scholes, G. D. Adv. Funct. Mater. 2008, 18, 1157–1172. (b) Sargent, E. H. Adv. Mater. 2005, 17, 515–522. (c) Eustis, S.; El-Sayed, M. A. Chem. Soc. Rev. 2006, 35, 209–217. (d) Salazar-Alvarez, G.; Qin, J.; S^epelak, V.; Bergmann, I.; Vasilakaki, M.; Trohidou, K. N.; Ardisson, J. D.; Macedo, W. A. A.; Mikhaylova, M.; Muhammed, M.; Baro, M. D.; Nogues, J. J. Am. Chem. Soc. 2008, 130, 13234–13239. (2) Innocenzi, P.; Kidchob, T.; Falcaro, P.; Takahashi, M. Chem. Mater. 2008, 20, 607–614. (3) (a) Prasad, B. L. V.; Sorensen, C. M.; Klabunde, K. J. Chem. Soc. Rev. 2008, 37, 1871–1883. (b) Joseph, S. T. S.; Itty Ipe, B.; Pramod, P.; Thomas, K. G. J. Phys. Chem. B 2006, 110, 150–157. (c) Sun, S. Adv. Mater. 2006, 18, 393–403. (d) Pileni, M. P. Acc. Chem. Res. 2007, 40, 685–693. (4) (a) Wang, D.; M€ohwald, H. J . Mater. Chem. 2004, 14, 459–468. (b) Xia, Y.; Gates, B.; Yin, Y.; Lu, Y. Adv. Mater. 2000, 12, 693–713. (c) Kraus, T.; Malaquin, L.; Delamarche, E.; Schmid, H.; Spencer, N. D.; Wolf, H. Adv. Mater. 2005, 17, 2438– 2442. (d) Malaquin, L.; Kraus, T.; Schmid, H.; Delamarche, E.; Wolf, H. Langmuir 2007, 23, 11513–11521. (5) Maury, P.; Escalante, M.; Reinhoudt, D. N.; Huskens, J. Adv. Mater. 2005, 17, 2718–2723. (6) (a) Tekin, E.; Smith, P. J.; Hoeppener, S.; Van den Berg, A. M. J.; Susha, A. S.; Rogach, A. L.; Feldmann, J.; Schubert, U. S. Adv. Funct. Mater. 2007, 17, 23–28. (b) Ko, H.-Y.; Park, J.; Shin, H.; Moon, J. Chem. Mater. 2004, 16, 4212–4215. (7) (a) Garcia, R.; Martinez, R. V.; Martinez, J. Chem. Soc. Rev. 2006, 35, 29–38. (b) Lee, K.-B.; Lim, J.-H.; Mirkin, C. A. J. Am. Chem. Soc. 2003, 125, 5588–5589. (c) Basnar, B.; Willner, I. Small 2009, 5, 28–44.

14294 DOI: 10.1021/la1023339

template-induced assembly,4 chemically driven assembly,5 and various printing techniques (inkjet printing,6 dip-pen nanolithography,7 soft lithography8). Efficient methodologies often combine topdown techniques for the fabrication of patterned substrates with bottom-up approaches for conveniently producing particles with complex structures.9 The patterning tasks are thus split into a more conventional top-down component, based on a well-established microfabrication technique, and on a bottom-up part, which takes advantage of colloidal phenomena, self-assembly, and novel building blocks. Templated assembly can allow for local particle positioning4-10 by exploiting a precise definition of the substrate topography and careful control of the nature of interactions among particles and between particles and the templating surface. This approach requires knowledge of phenomena such as sedimentation,11 gravitation, convective flux, capillary,4c,d,5-12 or centrifugal forces,13 which can further influence the overall process. For example, the assembly of bare submicrometer spherical beads, without inherent functionality, either inorganic or organic in nature (i.e., silica and polystyrene), into shallow hole patterns or grooves has been demonstrated.4 The challenge is now to convey optical, magnetic, and electrical properties to particle-based architectures and thus integrate these (8) (a) Lee, I.; Zheng, H.; Rubner, M. F.; Hammond, P. T. Adv. Mater. 2002, 14, 572–577. (b) Yan, X.; J.; Yao, J.; Lu, G.; Li, X.; Zhang, J.; Han, K.; Yang, B. J. Am. Chem. Soc. 2005, 127, 7688–7689. (9) Cheng, J. Y.; Ross, C. A.; Smith, H. I.; Thomas, E. L. Adv. Mater. 2006, 18, 2505–2521. (10) (a) Rycenga, M.; Camarago, C. P. H.; Xia, Y. Soft Matter 2009, 5, 1129– 1136. (b) Xia, Y.; Yin, Y.; Lu, Y.; McLellan, J. Adv. Funct. Mater. 2003, 13, 907–918. (11) (a) van Blaaderen, A.; Ruel, R.; Wiltzius, P. Nature 1997, 385, 321–324. (b) Dziomkina, N. V.; Vancso, G. J. Soft Matter 2005, 1, 265–279. (12) Kuncicky, D. M.; Prevo, B. G.; Velev, O. D. J. Mater. Chem. 2006, 16, 1207–1211. (13) Varghese, B.; Cheong, F. C.; Sindhu, S.; Yu, T.; Lim, C.- T.; Valiyaveettil, S.; Sow, C.-H. Langmuir 2006, 22, 8248–8252.

Published on Web 08/05/2010

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functions into devices. For this purpose, a precise design and an accurate control of the stability and processability of the particles are needed. The functionalization of single colloidal beads with magnetic, metal, and fluorescent nanocrystals (NCs) is one route to transfer the nanocrystals’ size- and shape-dependent properties to the final product. Recently, luminescence of semiconductor NCs has attracted increasing attention14 as novel fluorescent markers able to replace organic fluorophores thanks to their (i) size-dependent optical properties, (ii) narrow emission band, (iii) broad absorption band, and (iv) inherent robustness against quenching. NCs of different size can be excited with a single wavelength, and their narrow emission in the visible and near-IR allows multicolor detection, which makes these materials appealing for application in multiplex assays, bar coding, etc.15 Several strategies have been reported to label polystyrene (PS) beads with luminescent NCs, such as embedding hydrophobic NCs into polymer subsurface regions by solvent swelling and evaporation,16 incorporating NCs into the polymer precursor matrix, and subsequently fabricating functionalized beads by in situ polymerization17 or coating the bead surface with NCs. Among the various approaches, NC deposition onto coated beads treated with polyelectrolytes using a layer-by-layer (LbL) procedure18 represents a simple and reliable route. The relatively time-consuming multistep coating procedure is rewarded by excellent control of the NC and bead properties, i.e., the size-dependent properties of the NCs as well as the size monodispersity of the beads are preserved, which is crucial for the positioning procedure. Here we present a reliable strategy that combines the preparation of polymer beads functionalized with luminescent NCs and the capillary assembly technique for a precise positioning of each functional building block in suitable patterns. In particular, a LbL polyelectrolyte (PE) deposition procedure has been implemented that provides a highly uniform coverage of fluorescent NCs on PS beads. Polyelectrolyte-coated beads allow a more convenient and appropriate NC immobilization procedure than the single bare charged beads with a rougher and heterogeneously charged surface for further NC functionalization.19 Such decorated beads were positioned by templated-assisted capillary assembly onto patterned PDMS surfaces having arrays of holes and crosses in relief (Scheme 1).

Experimental Section Materials. Trioctylphosphine oxide (TOPO)/trioctylphine (TOP)-capped (CdSe)ZnS NCs were synthesized following the procedure reported in the literature with minor modifications.20b CdO powder, TOPO (technical grade 90%), TOP, hexadecylamine (HAD, 90%), tetrabutylphosphonic acid (TBPA, 97%), hexamethyldisilathiane (HMST), and diethylzinc (1 M solution in (14) (a) Somers, R. C.; Bawendi, M. G.; Nocera, D. G. Chem. Soc. Rev. 2007, 36, 579–591. (b) Zhou, M.; Ghosh, I. Peptide Sci. 2007, 88, 325–339. (15) (a) Battersby, B. J.; Lawrie, G. A.; Johnston, A. P. R.; Trau, M. Chem. Commun. 2002, 1435–1441. (b) Eastman, P. S.; Ruan, W.; Doctolero, M.; Nuttall, R.; De Feo, G.; Park, J. S.; Chu, J. S. F.; Cooke, P.; Gray, J. W.; Li, S.; Chen, F. F. Nano Lett. 2006, 6, 1059–1064. (16) (a) Gao, X.; Cui, Y.; Levenson, R. M.; Chung, L. W. K.; Nie, S. Nature Biotechnol. 2004, 22, 969–976. (b) Xu, C.; Bakker, E. Anal. Chem. 2007, 79, 3716– 3723. (c) Han, M.; Gao, X.; Su, J. Z.; Nie, S. Nature Biotechnol. 2001, 19, 631–635. (d) Sathe, T. R.; Agrawal, A.; Nie, S. Anal. Chem. 2006, 78, 5627–5632. (17) (a) Li, Y.; Liu, E. C. Y.; Pickett, N.; Skabara, P. J.; Cummins, S. S.; Ryley, S.; Sutherland, A. J.; O’Brien, P. J. Mater. Chem. 2005, 15, 1238–1243. (b) Sheng, W.; Kim, S.; Lee, J.; Kim, S.-W.; Jensen, K.; Bawendi, M. G. Langmuir 2006, 22, 3782–3790. (18) (a) Caruso, F.; Susha, A. S.; Giersig, M.; M€ohwald, H. Adv. Mater. 1999, 11, 950–953. (b) Ni Allen, C.; Lequeux, N.; Chassenieux, C.; Tessier, G.; Dubertret, B. Adv. Mater. 2007, 19, 4420–4425. (c) Li, J.; Zhao, X.-W.; Zhao, Y.-J.; Gu, Z.-Z. Chem. Commun. 2009, 2329–2331. (d) Sukhorukov, G.; Fery, A.; M€ohwald, H. Prog. Polym. Sci. 2005, 30, 885–897. (19) Wang, D.; Rogach, A. L.; Caruso, F. Nano Lett. 2002, 2, 857–861.

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Scheme 1. Template-Assisted Capillary Assembly of NC-Functionalized Polyelectrolyte-Coated Polystyrene Beads

hexane) were purchased from Sigma-Aldrich. Carboxylate-functionalized PS beads (d = 500 nm with a particle concentration 2.5 wt %) were purchased from Polymer Science. 11-Mercaptoundecanoic acid (MUA), tetrabutylammonium hydroxide, TBAH (1 M solution in methanol), poly(allylamine) hydrochloride (PAH, MW = 70 000), poly(diallyl)dimethylammonium chloride (PDDA, low molecular weight, 20% water), and poly(sodium 4-styrenesulfonate) (PSS, MW = 70 000) were purchased from Sigma-Aldrich. Polyelectrolyte solution were prepared by dissolving the polyelectrolyte powder in either PBS buffer or carbonate buffer, while the NC water solution was prepared using a carbonate buffer as solvent with [NaCl] = 0.5 or 0.05 M. For all experiments, Milli-Q water was used. All chemicals were used as received. Ligand-Exchange Reaction. 0.5 mL of the as-synthesized organic soluble TOPO/TOP-capped NCs (nearly 10-5 M) were diluted 1:3 with chloroform, and the new ligand MUA (10 mg/mL) was added. The solution was stirred overnight. It became turbid upon addition of a small amount of organic base (TBAH). At this stage, the nanocrystal solution was collected by centrifugation at 3000 rpm and dissolved in carbonate buffer. Repeated centrifugation and dialysis were carried out to remove the insoluble particles and the free ligand molecules, respectively, from the solution. UV-vis absorbance and PL spectra were recorded before and after the ligand exchange reaction, and relative quantum yields were calculated using Rhodamine 101 as reference.

Polyelectrolyte Coating of PS Beads and Their NC Functionalization. Stock PE solutions were prepared by dissolving PAH, PDDA, and PSS (10 mg/mL) in either a PBS buffer or a carbonate buffer at [NaCl] = 0.5 M. The polycation (PAH or PDDA) solution was added to the carboxylate-functionalized bead suspension (diluted to 60 μL/mL from the commercial batch, resulting in a final concentration of 2.7  1010 particles/mL). After 20 min of alternately stirring and sonication, centrifugation and final redispersion in buffer solution with reduced salt content ([NaCl] = 0.05M) were required for further characterization. The washing step is necessary to remove the free PE that could interfere with the next deposition step of the oppositely charged PE. This procedure was repeated for each layer in the multilayer structures (PEM), ending with an outermost positively charged layer. At this stage, MUA-capped (CdSe)ZnS water solution (10-6 M) was added to the PEM-coated PS beads, and the centrifugation/redispersion steps were carried out to remove unbound NCs. Finally, the last two outer PE layers were assembled to improve the colloidal stability. Characterization of NC and NC-Labeled PS Beads. UV-vis absorption measurements were acquired using a UV/vis/NIR Cary DOI: 10.1021/la1023339



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5 spectrophotometer (Varian). The luminescence spectra were recorded by means of the Eclypse spectrofluorimeter (Varian). The optical measurements on the NC solutions were performed at room temperature on the solution obtained directly from synthesis without any size-sorting steps and on the NC water solution after the ligand-exchange reaction. TEM analysis was performed using a Jeol JEM-1011 microscope operated at 100 kV. The samples were prepared by depositing dilute NC solutions onto 400-mesh carbon-coated copper grids and leaving the solvent to evaporate. TEM images of the nanocrystals before and after the phase transfer and of the PS beads at every stage of the assembly were recorded. The electrophoretic mobility of the diluted water bead suspensions with fixed salt content (0.05 M) was measured at each step of the multilayer fabrication using a ZetaPALS instrument (Brookhaven), usually employed for measurements on particles ranging from a few nanometers to as much as 30 μm diameter. The amplitude-weighted phase difference in the electrophoretic light scattering is used to determine particle mobility and, from this, the zeta potential value.22

Capillary-Assisted Template-Induced Bead Assembly.

The PDMS templates were fabricated as reported,4d using a patterned silicon substrate as master. In particular, the molded PDMS templates had arrays of (i) 200-nm-high cross-shaped features and (ii) holes of 2 μm diameter and 1-1.2 μm depth, with a hole density of 9  104 holes/mm2. The capillary assembly of the fluorescent NC-functionalized beads was carried out using a horizontal setup4c,d with precise control of the stage speed and temperature. The optical microscope images were acquired with a Zeiss Axiotech (Carl Zeiss AG, Germany), and the SEM images of the particle assemblies on the templates were recorded with a LEO 1550 SEM (Carl Zeiss AG, Germany).

Results and Discussion Nanocrystal Phase Transfer. Colloidal (CdSe)ZnS NC synthesis was carried out to provide luminescent inorganic markers to decorate carboxylate-functionalized PS beads. The functionalization of micrometer-sized PS particles needs to be carried out in aqueous media and, therefore, requires the availability of water-soluble NCs. However, the commonly used (CdSe)ZnS NC syntheses in hot coordinating solvent provide NCs soluble in organic solvents, with a higher control over particle size, monodispersity, and crystallinity. Therefore, the functionalization of PS beads with NCs require the use of postsynthetic strategies to ensure NC solubility in water and hence their subsequent processability. In particular, luminescent CdSe core NCs were synthesized by decomposition of organometallic precursors (CdO) in hot coordinating solvents,20 namely, TOPO, TOP, and HDA, followed by in situ ZnS shell growth. The synthesis yielded NCs soluble in organic solvent, coated with an organic aliphatic capping layer whose optical properties depend on the core size, and were preserved thanks to the ZnS shell that confines the charge carriers inside the core. In this work, following such a synthetic approach, two NC samples of different sizes were prepared. The two samples were characterized by UV-vis absorbance bands with a first excitonic peak centered at 564 and 587 nm, respectively (Figure 1A), which correspond to 3- and 4-nm-sized particles20b, and as confirmed by transmission electron microscopy (TEM) images (inset Figure 1A, bottom and top, respectively). The PL spectra of the related samples are reported in Figure 1B along with the picture (inset, Figure 1B) of the UV lamp illuminated NC solutions, which clearly highlight the NC (20) (a) Mekis, I.; Talapin, D. V.; Kornowski, A.; Haase, M.; Weller, H. J. Phys. Chem. B 2003, 107, 7454–7462. (b) Yu, W. W.; Qu, L.; Guo, W.; Peng, X. Chem. Mater. 2003, 15, 2854–2860.

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Figure 1. UV-vis absorbance spectra of 3 nm (straight line) and 4 nm diameter (dashed line) (CdSe)ZnS NCs in CHCl3 (A) together with the normalized photoluminescence (PL) spectra before (B) and after (C) NC surface processing and phase transfer. TEM images (inset A) of the NC samples (3-nm-sized NCs, inset A bottom; 4-nm-sized NCs, inset A top) and related picture of the NC solutions under UVlight lamp illumination (inset B). Quantum yield for 3- and 4-nm NC, right and left, respectively, before and after phase transfer (inset C).

size-dependent emission. Moreover, a narrow line width (129 and 120 meV for the 3- and 4-nm sized NCs) underlines the narrow size distribution as confirmed by TEM analysis (data not shown). The as-prepared organic soluble NCs were then prompt to be further transferred into aqueous media. Of the several approaches developed to prepare water-soluble (CdSe)ZnS NCs, ligand exchange reaction was an easy and straightforward strategy to follow. In particular, mercaptocarboxylic acids21 are interesting molecules for ligand exchange to achieve the phase transfer of hydrophobic-coated NCs into water. The strong coordination of the terminating thiol group with the zinc sulfide shell of the NCs can effectively replace the original TOPO/TOP ligands. Concomitantly, the carboxylic group added solubility in polar media, with a net negative surface charge of the NCs at basic pH. Mercaptoundecanoic acid (MUA) was selected because the relatively long carbon chain may be beneficial in preserving the NC photoluminescence (PL), but still allowing the NCs to be dissolved in water. The normalized PL spectra of water-soluble NC samples are reported in Figure 1C, showing that NCs retain their luminescence with only slightly decreased intensity and a minor red shift upon ligand exchange and transfer into the aqueous environment, which is still convenient for the emission detection. This modification in the emission features can be ascribed to a change of the dielectric constant of the solvent and/or to a NC surface chemistry modification. The relative PL quantum yields (QY) of NCs in CHCl3 and in water were calculated (using Rhodamine 101 as a standard) and reported in the inset of Figure 1C. Moreover, the nearly unchanged PL line width, corresponding to the samples before (129 and 120 meV for the 3- and 4-nm sized NCs) and after (120 and 123 meV) the ligand exchange reaction, highlights that (21) Chen, W. C. W.; Nie, S. Science 1998, 281, 2016–2018. (22) Villari, V.; Micali, N. J. Pharm. Sci. 2008, 97, 1703–1730.

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Figure 2. FT-IR spectra in ATR mode of TOPO/TOP-capped NCs (A), MUA (B), and MUA-capped NCs (C).

the size distribution does not broaden upon the phase transfer process. FTIR spectroscopy was used to probe the NC surface chemistry to monitor the effectiveness of the ligand exchange reaction. In Figure 2, FTIR spectra in ATR sampling mode of the TOPO/ TOP-capped (Figure 2A) and MUA-coated (Figure 2C) NCs are reported along with the neat MUA (Figure 2B). The NC samples after the surface-exchange reaction (Figure 2C) exhibit intense peaks at 2960 cm-1 ascribable to the antisymmetric stretching of the C-H of the methyl groups and at 2919 and 2850 cm-1 attributed to the symmetric and antisymmetric stretching of the C-H of the methylenic groups, respectively, arising from the hydrocarbon chains covering the NC surface. Moreover, a band centered at 1053 cm-1 is also clearly detected and inferred to belong to the characteristic PdO stretching mode of the phosphine oxide molecules, which have not been completely removed from the NC surface during the ligand-exchange reaction. In the same spectra, the presence of two distinguishable bands centered at 1583 and 1443 cm-1 is univocally attributed to the antisymmetric and symmetric stretching vibrations of carboxylate anions and confirms the effectiveness of the surface modification. Conversely, the disappearance of the intense -CdO stretching mode at 1703 cm-1 of the two bands centered at 1438 and 1260 cm-1, ascribable to the out-of-plane C-O-H bending, and to the C-O stretching of dimeric, and of the band at 939 cm-1, corresponding to the outof-plane O-H bending, which are all attributed to the carboxylic moiety of the free ligand, allows us to exclude the presence of unionized monomers, as expected at the pH value of the water solution. Moreover, the lower intense shoulder from 2700 to 2500 cm-1 associated with the S-H stretching of the mercaptocarboxylic acid (Figure 2B) is not still detected in the spectra of MUAfunctionalized NC (Figure 2C) because of the thiol deprotonation at high basic pH. NC stability against precipitation was also tested. To avoid NC flocculation, the pH of the colloidal solution was adjusted to a value higher than the pKa of the carboxylic moiety, thus the colloidal stability depends also on pH and not only on the concentration of carboxylate groups coordinating the Langmuir 2010, 26(17), 14294–14300


NC surface. In addition, the NC colloidal stability in aqueous media can be accounted for by the thiol moieties. Indeed, once dissociated as thiolate, at pH = 9, such functional groups provide a more effective coordination to the NC surface, thus protecting particle against flocculation. LbL Strategy toward the Fabrication of NC-Functionalized PS Beads. The hydrophilic luminescent NCs were then used to decorate positively charged PE-coated PS beads. First, the PE deposition parameters, i.e., PE composition and concentration,23 number of PE layers, ionic strength of the medium, and nanocrystalto-bead ratio, were systematically investigated. We found them to be key factors in controlling the fabrication of a stable colloidal suspension. Soft and charged spheres were thus built up, ready for functionalization with hydrophilic fluorescent NCs. Several factors influence the formation and stability of colloidal dispersions containing spherical particles surrounded by LbL polymeric structures prepared by electrostatic-force-induced deposition technique.23 In particular, the stability of colloidal particles against flocculation can be evaluated with respect to the polyelectrolyte concentration (C). Two PE limit concentrations can be considered: Csat, the PE concentration at which a complete particle coverage (saturation limit) is attained, and Cdep, the PE concentration at which the attractive depletion forces of the free PE in the bulk overcome the electrostatic repulsion. Accordingly, different regimes are reported to describe the colloidal stability of charged colloidal particles (Figure 3, upper panel). When C = 0, the bare particles are stable because the charge sites at the surface ensure electrostatic repulsion among the particles. Flocculation, instead, occurs for 0 < C < Csat, because if the PE content is lower than the saturation condition, it becomes statistically probable that two or more particles share the same PE unit. In this last regime, for C > Cdep, an excess of PE would result in a depletion attraction, able to overcome the electrostatic repulsion and, ultimately, trigger particle flocculation. It turns out that only in the PE concentration range Csat< C < Cdep the charged colloidal particles form a stable colloidal suspension. Moreover, also the saturation (τsat) and particle-toparticle collision time (τcol) must be taken into account in this regime, as flocculation can be avoided only if τcol > τsat. This means that the PE saturation at the particle surface has to be reached before particle collisions occur, thus inhibiting aggregation.23 Herein, PE solution with 10 mg/mL polymer content and [NaCl] at 0.5 M was found effective towards achieving a complete coverage of 500-nm-sized bead in the electrostatic-force-induced assembling procedure. A dilution of the salt (down to 0.05 M) was required during particle investigation to avoid the formation of crystallized NaCl residuals. The salt concentration is crucial for determining the ionic strength, and, consequently, the uniformity of the coating and the charge density along the PE chain in the multilayer structure. It is indeed known that the ionic strength affects the PE stiffness.24 For a fixed particle size, at high salt concentration, PEs assume a globular conformation, thus resulting in a thick coating of the colloidal particle. Conversely, at lower ionic strength, the polymer chains are known to arrange in a stretched conformation, thus resulting in an incomplete and not uniform assembly.24 In addition, at decreasing particle size, an increase in the salt content is required to reduce the polymer chain stiffness, so that a more uniform coverage at reduced curvature (23) (a) McCleents, J. Langmuir 2005, 21, 9777–9785. (b) Panchagnula, V.; Jeon, J.; Rusling, J. F.; Dobrynin, A. V. Langmuir 2005, 21, 1118–1125. (24) (a) Antipov, A. A.; Sukhorukov, G. B. Adv. Colloid Interface Sci. 2004, 111, 49–61. (b) Mayya, S. K.; Scoeler, B.; Caruso, F. Adv. Funct. Mater. 2003, 13, 80–84.

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Figure 3. Top: sketch of different colloidal stability regimes with respect to polyelectrolyte concentration. Bottom: ζ-potential evolution at each step of PS bead functionalization with PAH/PSS/PAH/NCs (A), PDDMA/PSS/PDDMA/NCs (B) at pH 7 at low (dashed line, B) and high (straight line, B) NC-to-PS bead ratio, and (PDDMA/PSS)2/PDDMA/NC-functionalized PS beads covered with two outer PE layers in carbonate buffer prepared by sonication (C).

radius is achieved. [NaCl] = 0.5 M was found to trade off these factors for the fabrication of polyelectrolyte multilayer structure on 500 nm bead. PAH, a weak, and PDDA, a strong polyelectrolyte,25 were alternatively used as polycation to be assembled onto carboxylatefunctionalized PS beads in a layer-by-layer structure, where PSS were used as polyanion. A weak polyelectrolyte charge density strongly depends on pH conditions; therefore, preliminary assembling experiments were carried out at pH = 7, to ensure a significant positive charge density along the PAH chain, without being detrimental to NC stability. In Figure 3, lower panel, the ζ-potential data are plotted versus the number of layers wrapped around the PS beads. In particular, Figure 3A,B shows the ζ-potential evolution corresponding to the preparation of PS beads coated with three layers of PAH/PSS/PAH (Figure 3, panel A) and with PDDA/PSS/PDDA (Figure 3, panel B). The last value of the five-point segment is referred to PS beads decorated with NCs. In both cases, positive and negative values of the ζ-potential alternate, marking the charge reversal of the surface occurring when polycation and polyanion or NC, respectively, covered the bead surface. Finally, NC deposition onto PDDA/PSS/PDDA-coated PS beads was studied and the effect of the NC concentration investigated. For this purpose, NC-to-PS bead ratios of 50 and 150 were tested. As reported in Figure 3B, a negative ζ-potential value is registered for NC-to-PS bead ratio around 150 (solid line), while a positive value is measured for the lower NC-to-PS ratio. The latter can be accounted for by an incomplete negatively charged NC coverage of the PE-functionalized positive bead surfaces. The morphology of NC-decorated PS beads at different particle ratios was also investigated by TEM analysis. In (25) Burke, S. E.; Berrett, C. J. Langmuir 2003, 19, 3297–3303.

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Figure 4. TEM images of three PE-coated PS beads (A) functionalized with NC at low (B) and high (C) NC-to-PS bead ratio prepared by stirring and sonication (D).

Figure 4, TEM images of the PE-coated PS beads (Figure 4A) and the NC-decorated PE-coated beads at low (Figure 4B) and high (Figure 4C) NC-to-PS bead ratios are reported. A low NC-to-PS bead ratio (≈ 50) results in an incomplete bead surface coverage (Figure 4B), while a rather uniform bead coverage is attained for NC-to-bead ratios around 100 (Figure 4C), thus confirming the ζ-potential conclusions. Finally, the NC-to-bead ratios of 150 was found to result only in an excess of NCs in the superrnatant upon centrifugation. Therefore, a value around 100 was selected to achieve complete bead coverage and to lessen aggregation phenomena occurring at low NC-to-PS-bead ratios (