Functional and Multifunctional Nanoparticles for Bioimaging and Biosensing

Functional and Multifunctional Nanoparticles for Bioimaging and Biosensing

(Parte 1 de 3)

DOI: 10.1021/la903512m ALangmuir X, X(X), X–X ©XXXX American Chemical Society


Subramanian Tamil Selvan,*,† Timothy Thatt Yang Tan,‡ Dong Kee Yi,§ and Nikhil R. Jana )

†InstituteofMaterialsResearchandEngineering,3ResearchLink,Singapore117602,‡DivisionofChemicaland Biomolecular Engineering, School ofChemical and Biomedical Engineering, NanyangTechnological University, 62 Nanyang Drive, Singapore 637459, §Division of Bionanotechnology, Kyungwon University, Seong Nam City,

Republic of Korea, and ) Centre for Advanced Materials, Indian Association for the Cultivation of Science,

Kolkata-700032, India

Received September 17, 2009. Revised Manuscript Received November 10, 2009

Herein, we describe the synthesis of functional and multifunctional nanoparticles (NPs), derived from our recent work, for bioimaging and biosensing applications. The functionalized NPs involve quantum dots (QDs), magnetic particles (MPs) and noble metal NPs for the aforementioned applications. A diverse silica coating approaches (reverse microemulsion and thin silanization) are delineated for the design of water-soluble NPs. We also review the synthesis of silica-coated bifunctional NPs consisting of MPs and QDs for live cell imaging of human liver cancer cells (HepG2) and mouse fibroblast cells (NIH-3T3). Using silica coated NPs, various NPs that are functionalized with antibody, oligonucleotide, biotin and dextran are efficiently used for protein detection.

1. Introduction

This feature article is organized as follows: (1) In the Introduction, we describe recent advances in inorganic nanoparticles (NPs) with more emphasis on quantum dots (QDs), magnetic NPs (MPs), and multifunctional NPs. (2) The significance of the coating is described for the creation of water-soluble functional NPs. (3) Silica coating approaches are delineated for the direct encapsulationofhydrophobicQDsandMPsorbifunctionalQDs and MPs within a silica shell using a reverse microemulsion method, derived from our recent work. We also describe a thin silica coating approach for the preparation of functional metallic NPs (e.g., Au and Ag), MPs, and QDs. (4) The functionalization ofNPswithvariouschemicalsandbiomoleculesarediscussed.(5) The biomedical applications of functional and multifunctional NPs with respect to live cell imaging and biosensingare described. (6) Finally, our conclusions and outlook are given.

Advances in Inorganic NPs. Inorganic NPs for biomedical applications have advanced rapidly in recent years because of their excellent optical and magnetic properties. An extensive amount of work has been done in the area of synthesis and surface modification of NPs.1 Semiconductor, noble metal, and metal oxide NPs of 1-100 nm have unique size-dependent properties, which prompted the researchers to investigate them in various imaging and sensing applications. The emerging applications of NPs in bioimaging and biosensing would lead to a variety of essential tools in medical diagnostics. Highly fluorescent semiconductor nanocrystals (NCs) or QDs (e.g., CdSe-ZnS) have emerged as a potential fluorescent label, owing to their remarkable optical properties.2 Compared to fluorescent dyes, QDs do not suffer the setback of photobleaching and their emission colors are tunable from the visible to the NIR region by varying the size or composition of QDs.

Recent advances in organometallic synthesis have enabled the size- and shape- dependent preparation of magnetic NPs

(MPs) (e.g., Fe2O3,F e3O4, Co, and FePt) and QDs (e.g., CdSe- ZnS, CdTe, and CdS) for applications in biology. A wide variety of contrast agents and optical labels are required for different types of imaging and detection modalities such as magnetic resonance imaging (MRI), positron emission tomography (PET), optical coherence tomography (OCT), and fluorescence-based imaging.3 Although each method has inherent advantages and drawbacks, MRI and PET are ideal for in vivo imaging, whereas fluorescence-based imaging is most widely employed for in vitro imaging.

Noble metal NPs such as gold (Au) and silver (Ag) are considered to be promising materials and are emerging as alternatives to semiconductor NCs.4 Noble metal NCs have surface plasmon resonance that induces a drastic enhancement in their absorption and scattering cross sections. This surface plasmon bandistunablefromthevisibletotheNIRregionbychangingthe particlesizeand shape from1-100 nmlengthscale.Theyarealso used asoptical-imaging-based contrast agents for the detection of proteins and DNA. Although individual particles of nanometer size can be imaged using scattered light or by the photothermal method, their application in bioimaging is largely limited because of background scattering from cellular organelles. Recent advances insyntheticmethodshavemadeitpossibletoexploreavariety of metallic nanostructures as optical contrast agents.5,6 Other types of magnetic oxide NPs have also been investigated for various biomedical applications.3,7 Superparamagnetic NPs are emergingasversatileprobesinbiomedicalapplications,especially in the area of MRI. Among these, iron oxide (γ-Fe2O3 or Fe3O4)

MPs have been widely used as T2 (negative) contrast agents. Additionally, manganese oxide (MnO) MPs are emerging as (1) Jun, Y.-W.; Lee, J.-H.; Cheon, J. Angew. Chem., Int. Ed. 2008, 47, 5122– 5135. (2) Medintz, I. L.; Uyeda, H. T.; Goldman, E. R.; Mattoussi, H. Nat. Mater. 2005, 4, 435–446.

(3) Weissleder, R. Science 2006, 312, 1168–1171. (4) Rosi, N. L.; Mirkin, C. A. Chem. Rev. 2005, 105, 1547–1562. (5) Jana, N. R. Small 2005, 1, 875–882. (6) Murphy, C. J.; Gole, A. M.; Stone, J. W.; Sisco, P. N.; Alkilani, A. M.;

Goldsmith, E. C.; Baxter, S. C. Acc. Chem. Res. 2008, 41, 1721–1730. (7) Huh, Y. M.; Jun, Y. W.; Song, H. T.; Kim, S.; Choi, J. S.; Lee, J. H.; Yoon,

S.; Kim, K. S.; Shin, J. S.; Suh, J. S.; Cheon, J. J. Am. Chem. Soc. 2005, 127, 12387– 12391.

B DOI: 10.1021/la903512m Langmuir X, X(X), X–X

Invited Feature Article Selvan et al.

promising contrast agents because of their size-dependent magnetic properties.7 These oxide MPs are also used in the magnetic harvesting of proteins and cells.

In clinical diagnosis, MRI is increasingly being used as an adjunct. To increase the tissue contrast in MRI applications, a number of contrast agents have been developed. Generally, the contrast agents are classified into two types: (a) positive and (b) negative. Positive and negative contrast agents are characterized by their ability to shorten either the longitudinal relaxation time,

T1, resulting in the brightening (hyperintense) of MR images, or the transversal relaxation time, T2, resulting in the darkening (hypointense) of MR images, respectively. The shortening of the relaxation time of water protons in the tissues dictates the utility and effectiveness of the contrast agents. The intrinsic T1 and T2 values of water are best described by their molar relaxivities, r1 and r2, which are expressed in mM-1 s-1. In clinical imaging, gadolinium-based paramagnetic chelates

(e.g., Gd-DTPA (DTPA is diethylene triamine pentaacetic acid)) are mainly used as T1 contrast agents.8 Introducing multifunctionalities such as fluorescence and a drug-targeting moiety onto

Gd-DTPA platforms has been found to be problematic. Conversely, iron oxide-based MPs could easily be modified to introduce a wide range of biological and pharmacological target- ing/deliverymoieties.Recently,MPs(Fe2O3,Fe3O4,andMFe2O4 where M=Ni, Co, Mn, or Fe) have been used as negative T2 contrast agents for MRI.9 Luminescent gadolinium oxide

(Gd2O3) hybrid MPs have been used as T1 contrast agents for both in vivo fluorescence and MR imaging.10 Nanoshells com- posed of Au3Cu1 have also been used as MR contrast agents.1 Advances in Multifunctional NPs. The fabrication of bi- functional or multifunctional NPs has received a great deal of attention in recent years because of their various biomedical applications.HybridinorganicNPsareemergingasusefulprobes for magnetic-based targeting, delivery, cell separation, MRI, and fluorescence-based biolabeling applications. Gold (Au) NPs have been extensively used to prepare multifunctional composites with

QDs and MPs. Typical examples include Fe3O4-Au,12

CdSe-Au,13 and PbSe-Au-Fe3O4.14 Multifunctional NPs have been actively explored for the en- hancement of imaging, targeting, and delivery. In the field of biological and biomedical imaging, QDs and MPs have been enjoyinggreater roles in biolabeling2 and MRI,15 respectively. A combinationofopticalandmagneticpropertiesinasinglematerial would enable simultaneous biolabeling/imaging and cell sorting/ separation.16,17 Nanocomposites consisting of semiconductor and magnetic NPs, known as magnetic quantum dots (MQDs),18 are of great interest as a new class of materials. Hybrid nanocompositessuch as Fe3O4-Au and CoPt-Au have shown the potential applications of such bifunctional NPs.19

More recently, silica-coated MPs with either QDs or dyes have beenusedintheaforementionedapplications.Wehavedeveloped silica coating methods for CdSe-ZnS20 and PbSe QDs21 and bifunctional NPs22,23 consisting of CdSe-ZnS QDs and γ-Fe2O3 MPs. Despite all of these advances, the application of multi- functional NPs in in vivo imaging is still in its infancy. In this feature article, we intend to discuss the synthesis and application of functional and multifunctional NPs derived from our recent work.

2. Functional and Multifunctional NPs: Significance of Coating

Water-soluble functional NPs are indispensable for various biomedical applications. However, the synthesis of robust functional NPs is very challenging because most of the good synthetic methods available for noble metal, QD, and magnetic oxides produce hydrophobic NPs as a result of the hydrophobic surfactant coating. Thus, water solubilization and functionalization are the key issues prior to their application, and herein lies the significance of the coating.24,25 The coating helps to convert hydrophobic NPs into hydrophilic water-soluble particles and introduce chemical functionality onto the particle surface so that different chemicals and biomolecules can be covalently attached. There are two common coating strategies that exist in order to convert hydrophobic NPs into hydrophilic and functional NPs. The first approach involves the ligand exchange of the original surfactant by hydrophilic ligands such as thiols or other functionalgroups.26Thiol-basedligandexchangeismostcommonfor noble metal NPs compared to other systems. This is because thiol brings about strong chemisorption on noble metal surfaces. A variety of thiol-based functional NPs of Au and Ag were synthesized.4 In addition, various approaches of thiol-based methods were developed to makea stable coating, whichinvolves theuseofligandswitheithermultiplethiols,thiolateddendrimers, dendrons, or the cross-linking of surface ligands.2,4

The second approach involves the interdigited bilayer formationbetweenamphiphilicmolecules/polymersandthepassivating surfactant layer surrounding NPs.27 These approaches have been successfully applied to noble metal NPs, in comparison with iron oxideMPsandQDs.Several methodsexistintheliteratureonthe design of water-soluble QDs.27-30 One method involves an

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(18) Ang, C. Y.; Giam, L.; Chan, Z. M.; Lin, A. W. H.; Gu, H.; Devlin, E.;

Papaefthymiou, G. C.; Selvan, S. T.; Ying, J. Y. Adv. Mater. 2009, 21, 869–873. (19) Cozzoli, P. D.; Pellegrino, T.; Manna, L. Chem. Soc. Rev. 2006, 35, 1195– 1208. (20) Selvan, S. T.; Tan, T. T.; Ying, J. Y. Adv. Mater. 2005, 17, 1620–1625. (21) Tan, T. T.; Selvan, S. T.; Zhao, L.; Gao, S.; Ying, J. Y. Chem. Mater. 2007, 19,3 112–3117. (2) Yi, D. K.; Selvan, S. T.; Lee, S. S.; Papaefthymiou, G. C.; Kundaliya, D.;

Ying, J. Y. J. Am. Chem. Soc. 2005, 127, 4990–4991. (23) Selvan, S. T.; Patra, P. K.; Ang, C. Y.; Ying, J. Y. Angew. Chem., Int. Ed. 2007, 46, 2448–2452. (24) Gerion, D.; Pinaud, F.; Williams, S. C.; Parak, W. J.; Zanchet, D.; Weiss,

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DOI: 10.1021/la903512m CLangmuir X, X(X), X–X

Selvan et al. Invited Feature Article organic coating using either polymers,28 micelles,29 or thiols30 as the linker molecules.Another method isbased onthe well-known silica chemistry developed for coating metal NPs.31

Significant research effort has been devoted to preparing core-shell-type NPs with a cross-linked shell that could protect the NPs much better than a thiol-based coating. The most widely used approach is a silica coating24 along with other methods such as ligand or polymer bridging25 and the incorporation of NPs within microparticles.32 Although a variety of functional NPs havebeensynthesized bya silicaorpolymercoating, eachcoating methodhasinherentadvantagesandlimitations.Itwouldbeideal to have a thin, cross-linked coating that could protect the core, improve colloidal stability, and introduce chemical functionality forbioconjugation.Effortsarestillunderwayinvariousgroupsto makea library ofrobustfunctional NPs. Inthisfeature article,we will focus on silica coating approaches developed by us for creating various robust functional NPs. To utilize functional NPs for biomedical imaging, one has to focus on the following steps: (a) synthesis, (b) coating, (c) surface functionalization or bioconjugation, and (d) applications, as depicted in Scheme 1.

3. Silica Coating

(Parte 1 de 3)