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Fluorescent and paramagnetic coreshell hybrid nanoparticles for bi-modal

magnetic resonance/luminescence imaging

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Published on 14 August 2012 on http://pubs.rsc.org | doi:10.1039/C2JM34508K

Weihua Di,*a Sabareesh K. P. Velu,bcd Alessandro Lascialfari,bcd Chunxu Liu,e Nicola Pinna,fg Paolo Arosio,b
Yoshio Sakka*hi and Weiping Qin*a
Received 10th July 2012, Accepted 14th August 2012
DOI: 10.1039/c2jm34508k
Well calibrated coreshell multifunctional nanoparticles for biomedical applications were synthesized
by a multistep soft chemistry route. The core is composed of Gd(OH)CO3$H2O spheres prepared via a
urea-based homogeneous precipitation technique, while the shell is a homogeneous thin silica layer
embedded with the fluorescent dye rhodamine B (RhB) prepared via a modified St
ober process. The
hybrid coreshell nanoparticles show a paramagnetic behavior with a specific saturation magnetization
of 2.8 emu g
1. The nuclear magnetic resonance relaxation measurements reveal that these systems
could be used as T1 and T2 magnetic resonance imaging (MRI) contrast agents. Also, the resulting
coreshell nanoparticles are fluorescent due to the presence of RhB entrapped inside the silica shell.
When incubated with the human cervical carcinoma (HeLa) cells the coreshell composite particles
exhibit bright intracellular fluorescence, indicating their capability for optical imaging in biology.
Furthermore, the incorporation of organic dyes inside the silica matrix yields outstanding advantages
such as significantly improved photostability of the dye and reduced cytotoxicity due to the protection
of biocompatible silica shell. These features demonstrate that the magnetofluorescent coreshell
nanoparticles prepared in our work have the potential to serve as a versatile imaging tool for smart
detection or diagnosis in future biomedical engineering.

1. Introduction
The application of nanomaterials in medical and biological fields
has drawn great research interest in recent years.14 Currently,
a
State Key Laboratory on Integrated Optoelectronics, College of Electronic
Science and Engineering, Jilin University, Changchun 130012, P.R. China.
E-mail: whdi@jlu.edu.cn, wpqin@jlu.edu.cn; Fax: +86-431-851682408325; Tel: +86-431-85168240-8325
b
Dipartimento di Fisica, Universit
a degli Studi di Milano, Via Celoria 16,
Milano 20133, Italy
c
Dipartimento di Fisica A.Volta, Universit
a degli studi di Pavia, Pavia
27100, Italy
d
CNR-Istituto di Nanoscienze-S3, Modena, Italy
e
State Key Laboratory of Luminescence and Application, Changchun
Institute of Optics, Fine Mechanics and Physics, Chinese Academy of
Sciences, Changchun 130033, P.R. China
f
Department of Chemistry, CICECO, University of Aveiro, 3810-193
Aveiro, Portugal
g
World Class University (WCU) program of Chemical Convergence for
Energy & Environment (C2E2), School of Chemical and Biological
Engineering, College of Engineering, Seoul National University (SNU),
Seoul 151-744, Korea
h
Materials Processing Unit, National Institute for Materials Science, 1-2-1
Sengen, Tsukuba, Ibaraki 305-0047, Japan. E-mail: sakka.yoshio@nims.
go.jp
i
Graduate School of Pure and Applied Sciences, University of Tsukuba, 1-21 Sengen, Tsukuba, Ibaraki 305-0047, Japan
Electronic supplementary information (ESI) available: See DOI:
10.1039/c2jm34508k

This journal is The Royal Society of Chemistry 2012

nanoparticles have been used in many biological fields, such as

sensors,5,6 in vitro and in vivo fluorescent markers,7,8 clinical
diagnosis,9,10 drug delivery,11,12 and magnetic resonance imaging
(MRI) contrast agents.13,14
One of the critical challenges in nanobiotechnology is the
development of multifunctional particles at the nanoscale that
simultaneously fulfill different functionalities. The formation of
heterostructures with hierarchical morphologies can fulfill this
requirement due to the possibility of finely tuning the properties.15,16 Therefore, the synthesis of heterostructured nanoparticles
has attracted considerable attention in materials chemistry and
nanotechnology. In particular, coreshell nanoparticles can
combine diverse functionalities within a single nanoobject.1720
Coating the particles with a thin shell of a compatible material
permits control of the interparticle and particlematrix interactions, thereby further improving the functional properties and
expanding the range of potential applications. The structure, size
and composition of coreshell materials can be altered in a
controlled way in order to tailor their magnetic, optical, mechanical, thermal, electrical, sensing and catalytic properties.2026
For biological applications, functional nanomaterials must
also be biocompatible. For effective early cancer diagnosis and
treatment, the particles are required to be less than 200 nm in
diameter2729 and monodisperse to achieve efficient distribution in
the targeted tumor lesions.29 It should also be noted that
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macrophages tend to take up nanoparticles of diameters larger

than approximately 40 nm, thus reducing substantially the
possibility of reaching a specific target through the blood vessels.
In terms of shape, the spherical shape is preferred as it facilitates a
uniform surface conjugation of specific ligands (e.g., antibodies)
of the nanoparticle with the targeted cell.29 Long-wavelength
excitation (i.e. in the visible or near-infrared range) is also required
for deep tissue penetration for in vivo imaging.3032 In addition, the
combination of multiple functions into a nanoobject is still an
urgent need. For example, the introduction of fluorescent groups/
particles into drug carriers not only offers the ability to track/
visualize the targeted objects, but also provides real-time monitoring of the drug delivery behavior.11,33 Another example is the
incorporation of magnetic properties into fluorescent particles,
which not only allows multimodal imaging (e.g. magnetic resonance and fluorescence imaging),17,34,35 but also allows intracellular movements to be controlled using a magnetic field and
simultaneous monitoring using fluorescent microscopy.36
Based on these considerations, this work reports on the design
of well calibrated and uniform coreshell nanoparticles which are
simultaneously paramagnetic and fluorescent for bi-modal
magnetic resonance/fluorescence imaging monitoring.

2. Experimental
2.1.

Preparation of Gd(OH)CO3$H2O colloidal nanoparticles

The Gd(OH)CO3$H2O colloidal nanoparticles were prepared via

a urea-based homogeneous precipitation process.37,38 A total of
5 mL of Gd(NO3)3 (1 M) and 10 g of urea [CO(NH2)2] were
dissolved in deionized water. The total volume of the solution
was about 250 mL. The above solution was first homogenized
under magnetic stirring at room temperature for 2 h. The
resultant solution was then reacted at 85  C for 2.5 h in an oil
bath. The obtained suspension was separated by centrifugation
and collected after washing with deionized water several times.
2.2. Preparation of coreshell Gd(OH)CO3$H2O@SiO2
entrapped with rhodamine B (RhB)
The coreshell Gd(OH)CO3$H2O@SiO2 particles embedded
with RhB were prepared via a modified solgel process. In a
typical procedure, 50 mg of obtained spherical Gd(OH)
CO3$H2O particles and 3 mL of 2.5  10
4 M RhB solution were
well dispersed in a mixture of 70 mL of polyoxyethylene nonylphenyl ether, 125 mL of cyclohexane and 20 mL of deionized
water by ultrasonification for 20 min. 0.03 g of tetraethoxysilane
(TEOS) in 10 mL ethanol were added dropwise to the above
solution, followed by 1.0 mL of concentrated ammonia aqueous
solution (28 wt%). The solution was continuously stirred at room
temperature for 24 h. The above process was performed in the
dark. The obtained particles were separated and washed with
ethanol and water several times, and then dried in vacuum at
room temperature.
2.3.

Cell culture

HeLa cell lines were maintained in Dulbeccos modified Eagles

medium (DMEM) containing 10% fetal bovine serum, penicillin
(100 units mL
1), and streptomycin (100 mg mL
1). Cells were
20642 | J. Mater. Chem., 2012, 22, 2064120648

cultured with the complete medium in 5% CO2 at 37  C. For all

experiments, cells were harvested from subconfluent cultures by
the use of trypsin and were resuspended in fresh complete
medium before plating.
2.4.

Cell uptake and observation

In a typical procedure, 7.5  104 cells were plated in a 35 mm

petri dish for 4 h to allow the cells to attach. After the cells were
washed twice with phosphate buffered saline (PBS), the RhB
entrapped coreshell structured particles dispersed in PBS were
added to the petri dishes. After incubation for a period of time
(472 h), the cells were washed several times with PBS to remove
the remaining particles and dead cells, and then observed under
laser scanning confocal microscope (TCS SP5, Leica), operating
at around a 540 nm excitation wavelength using a tunable
Chameleon XR laser system.
2.5.

Cell viability and proliferation assay

In vitro cytotoxicity was assessed using the trypan blue exclusion

assay, HeLa cells were seeded at a concentration of 104 cells
mL
1 in a 12-well plate with 2 mL of culture medium per well.
The plate was incubated in 5% CO2 at 37  C. After 24 h, the
particles were added to the wells. A culture medium without
particles was used as the control. Cell numbers were measured
24, 48 and 72 h after adding the particles. The viability of
untreated cells was assumed to be 100%. The cytotoxicity was
expressed as the percentage of cell viability in the presence of
foreign materials compared to that of untreated control cells.
2.6.

Chemico-physical characterization

The transmission electron microscopy (TEM) observations were

carried out using a JEOL 2200FS microscope. Samples for TEM
investigations were prepared by first dispersing the particles in
ethanol with the aid of ultrasonification and then dropping 1
drop of the suspension onto a copper TEM grid coated with a
holey carbon film. The size and size distribution of the nanoparticles were extracted from TEM images based on the analysis
of 100 spheres which were randomly selected. Fourier transform
infrared (FT-IR) spectra (Mattson 5000) of the samples were
measured in the range of 4000500 cm
1 in transmission mode.
The pellets were prepared by adding 0.8 mg of the sample powder
to 80 mg of KBr. The magnetization curves were recorded on a
vibration sample magnetometer (VSM) (MOLSPIN, Newcastle
Upon Tyne, U.K.) at room temperature. For nuclear magnetic
resonance (NMR) relaxometry, the NMR longitudinal and
transverse relaxivities were collected at room temperature using
Stelar Spinmaster relaxometer in the frequency range 8 < n <
60 MHz. Standard radio frequency excitation pulse sequences
Carr-Purcell-Meiboom-Gill (CPMG) and saturation-recovery
were used to measure longitudinal (spin-lattice, T1) and transverse (spinspin, T2) relaxation times, respectively. The fluorescence spectra were recorded at room temperature with an
excitation wavelength of 540 nm on a fluorescence spectrophotometer (F-7000, Hitachi). The slit widths of the excitation and
emission were both 2.5 nm. For the measurement of fluorescence
evolution with time of pure RhB and RhB entrapped particles,
the fluorescence was monitored once every minute and a fast scan
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rate of 2400 nm min
1 was used. The Gd3+ concentration in the

sample was determined by an inductively coupled plasma atomic
emission spectrometer (ICP-AES) ICPS-8000 of the Shimadzu
Co. (Kyoto, Japan). The quantities of RhB on the adsorbed and
entrapped nanoparticles were roughly evaluated by comparing
the absorption of the supernatant after reaction with that of
original RhB solution.

3. Results and discussion

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3.1.

Gd(OH)CO3$H2O nanoparticles

The gadolinium hydroxylcarbonate (Gd(OH)CO3$H2O) was

prepared via a urea-based homogeneous precipitation method.
Urea serves as a precipitation agent due to self decomposition into
the OH
and CO32
at elevated temperatures (>83  C) providing
the anions for the precipitation of Gd(OH)CO3$H2O.37,38 Under
the TEM, the as-prepared particles appear spherical and nearly
monodispersed (Fig. 1(a)). The size distribution plot obtained from
100 particles selected randomly (Fig. 1(b)) indicates that the
nanoparticles range in size from 130 to 170 nm with an average
diameter of 150 nm and a standard deviation of 8.6%. The slow
and homogeneous release of the precipitating ligands (OH
and
CO32
) into the reaction system prevents inhomogeneous distribution of the reactants and allows precise control over nucleation
and growth.39 The size and shape of the particles obtained via the
urea-based precipitation process have been shown to depend on the
reaction conditions (the metal element, urea concentration, reaction time and so on).37,38,40,41 Lechevallier et al. recently reported
that an increase in reaction time, while the other conditions are kept
unchanged, leads to a transition of the morphology from spherical
to platelet-shaped particles.41 Chemical analysis of the supernatant
after removal of solid particles by centrifugation shows that the
concentration of Gd is below the detection limit of 0.01 wt% after
2.5 h of reaction at 85  C, indicating that the precipitation reaction
was complete. The as-prepared material is amorphous based on Xray powder diffraction measurements (not shown). The detailed
characterization of the as-synthesized Gd(OH)CO3$H2O nanoparticles is described in the ESI (Fig. S1, S2 and Table S1).

process, as described in the experimental section. The coreshell

structure of the particles after coating can be clearly seen by
TEM due to the different electron contrast of the cores and shells
(Fig. 2(a)(c)). The dark inner and gray outer regions correspond
to the Gd(OH)CO3$H2O core and SiO2 shell, respectively. It can
be concluded that the solgel process allows for a homogeneous
and complete coating around each spherical Gd(OH)CO3$H2O
particle and no separated SiO2 particles were formed. The
particles maintain spherical and monodisperse morphology after
coating.
The diameter of the coated particles shows a slight increase of
about 20 nm due to the homogeneous formation of about 10 nmthick SiO2 shell around the Gd(OH)CO3$H2O particles (Fig. 2
(c)). Similarly, the average particle size was measured from 100
coreshell particles randomly selected from TEM observations.
The size distribution plot (Fig. 2(d)) indicates that the nanoparticles range in size from 150 to 190 nm with an average
diameter of 170 nm and a standard deviation of 7.9%. The
standard deviation of particle size after coating is almost the
same as that before coating. This clearly indicates that the SiO2
coating is uniform. The easy growth of the homogeneous silica
shell on a Gd(OH)CO3$H2O particle could be attributed to the
composition of the Gd(OH)CO3$H2O particles. As a matter of
fact, due to the hydrated phase and the presence of hydroxyl
groups of as-prepared Gd(OH)CO3$H2O particles, TEOS can
easily react with the surface of the core particles, forming a first
silica monolayer on top of which the silica shell can easily grow.
The photoluminescence characterization of coreshell particles
provides the evidence that the RhB molecules are successfully
embedded into the SiO2 shell (see below).
3.3.

Photoluminescence properties

A drop of the coreshell particles dispersed in ethanol was

smeared on the surface of a glass slide for observation using a laser
scanning confocal microscope. Fig. 3 shows a fluorescence image

3.2. Coreshell structured Gd(OH)CO3$H2O@SiO2 particles

embedded with RhB
The spherical Gd(OH)CO3$H2O particle were coated by a
uniform layer of SiO2 embedded with RhB via a modified solgel

Fig. 1 TEM images of spherical Gd(OH)CO3$H2O particles (a) and the

size distribution plot based on the analysis of 100 particles from TEM
observations (b).

This journal is The Royal Society of Chemistry 2012

Fig. 2 TEM images of coreshell spherical particles (a, b and c) and the
size distribution plot based on the analysis of 100 particles from TEM
observations (d).

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of the coreshell particles upon optical excitation at the wavelength matching the absorption of RhB (450600 nm).42 The
particles are clearly visible due to the red emission of RhB
embedded in the SiO2 shell and appear uniformly dispersed on the
glass slide. This suggests that the Gd(OH)CO3$H2O@SiO2 core
shell nanoparticles are well dispersed and that the RhB dye could
be entrapped inside the SiO2 shell via the modified solgel process.
The emission behavior of the RhB doped coreshell particles was
further analyzed by laser photoluminescence spectroscopy at
room temperature. Upon excitation at 540 nm, the coreshell
particles exhibit a broad band emission centered at 582 nm, which
is the characteristic emission of RhB (Fig. 4). The emission of RhB
entrapped nanoparticles shows a slight blue-shift in comparison
with that of pure RhB. This is attributed to the reduction of
aggregation degree of RhB molecules in SiO2 matrix, compared
with that of free RhB molecules.43 Further, the emission quantum
yield of RhB entrapped nanoparticles is five times higher than that
of pure RhB. This is also attributed to a relatively homogeneous
distribution of RhB molecules in SiO2 matrix which reduces the
intermolecular luminescence quenching of RhB.43 A number of
studies on silica particles embedded with organic dyes by using the
St
ober or microemulsion method are available. Different types of
organic and metallorganic fluorophores, such as fluorescein isothiocyanate (FITC),44,45 rhodamine 6G (R6G),46 pyrene,47
RuII(bpy)3,48,49 and indocarbocyanine dyes,50 have been
successfully entrapped inside a silica matrix through physical or
chemical interactions.
The photostability measurements further confirm that RhB
molecules are entrapped inside the SiO2 shell. To check the
photostability of Gd(OH)CO3$H2O@SiO2 coreshell particles
entrapped with RhB, the relative fluorescence intensity as a
function of illumination time was monitored under a constant
illumination of 540 nm in wavelength. Only a slight decrease of
the photoluminescence intensity (less than 5% after 15 min) was
detected in the case of the RhB doped coreshell particles, suggesting a low degree of photobleaching. However, the free RhB
and the RhB adsorbed onto the Gd(OH)CO3$H2O@SiO2
particle surface show significant photobleaching with increased
irradiation time. As shown in Fig. 5, the relative photoluminescence intensity decreases to 38% and 45% of the initial

Fig. 3 Confocal fluorescence microscopy image of RhB embedded core

shell particles first dispersed in ethanol and then smeared on a glass slide.

20644 | J. Mater. Chem., 2012, 22, 2064120648

Fig. 4 Emission spectra of pure RhB (full line) and RhB entrapped
Gd(OH)CO3$H2O@SiO2 coreshell particles (dotted line) at room
temperature.

value upon 15 min illumination for the free RhB and the RhB
adsorbed onto the particle surface, respectively. These results
further demonstrate that the RhB molecules could be entrapped
inside the silica network via a modified solgel process and that
the photostability of RhB entrapped inside the silica network is
greatly increased compared to that of the dye itself. The
improved photostability is attributed to the caging effect,
which prevents or greatly reduces the contact between the dye
and molecular oxygen which is responsible for the photooxidation of organic dyes.51,52
3.4.

Magnetic properties

The magnetic properties of coreshell Gd(OH)CO3$H2O@SiO2

particles were studied using a vibration sample magnetometer
(VSM). Fig. 6 gives the room-temperature magnetization data of
the sample, indicating a paramagnetic behavior of coreshell
particles with a specific saturation magnetization of 2.8 emu g
1
due to the presence of Gd ions. Gd3+ ions have isotropic electronic ground state 8S7/2 and a half-filled f-orbital with seven
electrons and possess a high magnetic moment. Consequently,

Fig. 5 The photoluminescence intensity of pure RhB (triangles), RhB

adsorbed (circles) and entrapped (squares) coreshell particles against
illumination time.

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Fig. 6 The magnetization curves of Gd(OH)CO3$H2O@SiO2 particles

at room temperature.

the electronic relaxation time, T, is relatively long, 105106 times

higher than many other paramagnetic lanthanide ions.53 This
leads to large effects on both longitudinal and transverse proton
relaxivity of Gd3+ even at low applied magnetic fields of 1.5 T,
resulting in MRI contrast applicability for Gd3+ based materials.
Carboxylate complexes of Gd3+ are widely used as MRI contrast
agents in clinical practice.54 Gd3+DTPA (diethylene triamine
pentaacetic acid) complexes are used in clinics.55
The 1H NMR longitudinal and transverse relaxation times
were measured for coreshell Gd(OH)CO3$H2O@SiO2 particles
at room temperature in the frequency range 8 # n # 60 MHz.
This range of frequency was chosen to cover most of the clinical
frequencies (H 0.18 T
1.4 T).56 The efficiency of the contrast
agents is calculated from the nuclear relaxivity, (r), defined as the
inverse of the relaxation time normalized for contrast agent
concentration, once previously corrected for the host diamagnetic contribution, which is given by
ri [(1/Ti)meas
(1/Ti)dia]/c, i 1,2
where (1/Ti)meas is the measured value on the sample with
concentration c (mmol L
1) of magnetic center (in our case, [Gd3+]
1.835 mmol L
1) and (1/Ti)dia refers to the nuclear relaxation rate
of the diamagnetic host solution (water, in our case).57,58
Fig. 7 shows the r1 and r2 nuclear relaxivities of coreshell
Gd(OH)CO3$H2O@SiO2 particles together with the values for
the commercial contrast agents for r1 and r2 respectively. As can
be seen, the r1 values obtained for coreshell Gd(OH)
CO3$H2O@SiO2 particles are not far from to those of Omniscan
(commercial T1-relaxing paramagnetic contrast agent). Interestingly, the r2 values of coreshell Gd(OH)CO3$H2O@SiO2 particles are higher than those of Omniscan and are somewhat similar
to Endorem (T2-relaxing superparamagnetic contrast agents,
typical materials used for negative contrast).5862 Thus we
conclude that coreshell Gd(OH)CO3$H2O@SiO2 samples can be
considered as possible T1- and T2- relaxing MRI contrast agents.
3.5.

Cell imaging and cytotoxicity

To check the possibility for the use of RhB entrapped coreshell

particles as luminescent biological labels, we conducted in vitro
This journal is The Royal Society of Chemistry 2012

Fig. 7 Proton NMR longitudinal (a) and transverse (b) relaxivities of

Gd(OH)CO3$H2O@SiO2 particles measured at room temperature.

biological experiments using HeLa cells. A laser scanning

confocal microscope operating at around a 540 nm excitation
wavelength was applied to excite the RhB entrapped particles for
luminescence. As a control experiment, HeLa cells alone show
almost no background fluorescence upon excitation at 540 nm
(Fig. 8(a) and 8(b) correspond to the bright-field optical image of
HeLa cells alone). Nonetheless, upon incubation, incorporation
of RhB entrapped coreshell particles into HeLa cells was
observed. Fig. 8(c) and (d) show bright-field optical and fluorescence microscopy images of HeLa cells after incubation with
0.2 mg mL
1 of particles for 24 h, respectively. As observed, the
nanoparticles can permeate into the cell membrane and clearly
retain their intrinsic fluorescence upon cellular internalization
(Fig. 8(d)). Moreover, the corresponding bright-field measurements taken after incubation with the coreshell particles
confirmed that most of cells were viable throughout the imaging
experiments (Fig. 8(c)). To check whether the coreshell nanoparticles are internalized within the cells, we took a series of Zstack images of the cell (e.g. top to bottom) at 1 mm slice
intervals of the HeLa cells stained with coreshell nanoparticles.
A representative fluorescence image taken from an internal slice
(Fig. S4, ESI) clearly shows red fluorescence within the cells. It
can be reasonably concluded that the coreshell nanoparticles
are localized within the interior of the HeLa cells. It has been
reported that many kinds of inorganic or metal nanoparticles
with various shapes are able to be readily internalized by various
cancer cell lines.6368 However, the mechanism of nanoparticle
internalization has not been clarified precisely. It is now generally
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Fig. 8 Bright-field and fluorescence microscopy images of HeLa cells

incubated with RhB embedded coreshell particles for 48 h (c and d).
Similar images for HeLa cells without nanoparticles are also shown as a
control experiment (a and b).

accepted that serum proteins from biological media (e.g., bovine

serum albumin (BSA)) can nonspecifically coat the surfaces of
nanoparticles leading to modification of the effective surface
charge of the nanoparticles, regardless of their initial surface
charge.68,69 On this basis, the surface adsorbed BSA can facilitate
the uptake of particles into human cancer cells, such as HeLa
cells, via receptor-mediated endocytosis arising from cellular
recognition of these proteins. Certainly, the receptor-mediated
endocytosis mechanism can also explain our case, since the
culture media for HeLa cell lines in our experiment contains 10%
fetal bovine serum.
The cytotoxicity and biocompatibility of the RhB entrapped
coreshell particles were evaluated in order to further demonstrate their applicability in nanobiotechnology. The tests were
conducted following the trypan blue exclusion assay in order to
determine the number of viable cells. Fig. 9(a) shows the
concentration effect of RhB entrapped coreshell particles on cell
viability after incubation for 24 h. It demonstrates that the
particles show almost no toxicity upon incubation with HeLa
cells. As shown, the cell viability remains as high as 94% even
after incubation with a relatively high concentration of particles
(0.2 mg mL
1) for 24 h.
For biological applications, evaluation of the intermediate or
long term toxicity of foreign materials is also important. Fig. 9(b)
demonstrates the cell proliferation activity after incubation of
RhB entrapped coreshell particles of 0.2 mg mL
1with HeLa
cells for 24, 48 and 72 h, respectively. The number of HeLa cell
treated with the particles is increased rapidly, and the growth rate
of treated cells is similar to that of untreated cells. Even with an
incubation time of 72 h, the viability of the cells treated with
particles is still as high as 90%, relative to that of untreated cells
(Fig. 9(b)). In addition, the HeLa cells incubated with RhB
entrapped particles for various time were imaged by confocal
fluorescence microscopy upon excitation at 540 nm (Fig. 10). The
cells are clearly visible due to the bright red emission. From these
images, the number of HeLa cells increases rapidly with incubation time. For comparison, cells without incubation with
20646 | J. Mater. Chem., 2012, 22, 2064120648

Fig. 9 Concentration effect of RhB entrapped coreshell particles on

cell viability (a); the viabilities of HeLa cells incubated without and with
the coreshell particles as a function of time (b).

particles were optically imaged (Fig. S5, ESI). We can see a

similar cell growth trend as that of cells incubated in the presence
of particles. This indicates that cell proliferation was not
hindered significantly when HeLa cells were exposed to RhB
entrapped coreshell structured particles, further highlighting a
very low toxicity of the coreshell particles synthesized in this
work.

Fig. 10 Confocal fluorescence microscopy images of HeLa cells incubated with RhB embedded coreshell particles as a function of time. (a) 4
h; (b) 24 h; (c) 48 h; (d) 72 h.

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The toxicity of Gd(OH)CO3$H2O@SiO2 particles without

RhB entrapped, but with adsorption of RhB at the particle
surface was also measured. The viability of HeLa cells was 90%,
83%, and 72%, while incubated with the particles adsorbed with
RhB for 24, 48 and 72 h, respectively, indicating an increased
toxicity relatively to that of RhB entrapped particles. In our
previous work, we also conducted a cytotoxic evaluation for the
CePO4 : Tb nanorods adsorbed with and without RhB molecules at the particle surface. A relatively high toxicity of
CePO4 : Tb nanorods adsorbed with RhB molecules was
detected when they were incubated with HeLa cells for 72 h (cell
viability: 72%), but the bare CePO4 : Tb nanorods showed
almost no toxicity towards HeLa cells. It is well known that
organic dyes are often toxic, and thus the direct exposure of dye
molecules to cells can lead to a relatively high cytotoxicity. In
the present work, the RhB entrapped inside the silica matrix
prevents or greatly reduces the direct contact between RhB
molecules and the cells, thereby lowering the toxicity to cells as
well as improving the photostability upon irradiation, as
demonstrated above.

4. Conclusions
Well calibrated and uniform Gd(OH)CO3$H2O nanoparticles
were prepared using a urea-based precipitation process, and then
coated with a thin homogeneous layer of SiO2 entrapped with
fluorescent dye RhB, via a modified St
ober process, leading to the
formation of coreshell Gd(OH)CO3$H2O@SiO2 embedded
with RhB. The resulting coreshell particles endow the material
with both paramagnetic and fluorescence properties due to the
presence of paramagnetic gadolinium in the core and a fluorescent dye embedded in the silica shell, respectively. The potential
of coreshell particles for use as contrast agents in MRI applications was demonstrated by the proton NMR longitudinal r1
and transverse r2 relaxivities that are not far from (r1) and higher
than (r2) those of commercial paramagnetic (positive) contrast
agents. Upon incubation with HeLa cells, the coreshell nanoparticles can permeate into the cell membrane and clearly show
their intrinsic fluorescence upon cellular internalization, suggesting their potential for cell imaging. The incorporation of
RhB dyes inside the silica matrix improved the photostability
and reduced the cytotoxicity significantly, compared to that of
free dye. This work demonstrates that the paramagnetic and
fluorescent coreshell nanoparticles are promising candidates for
magnetic resonance and photoluminescence dual-mode imaging
applications.

Acknowledgements
This study was partly supported by the National Natural Science
Foundation of China (Grant nos. 61178073), the Grant-in-Aid
for Scientific Research of the JSPS and the World Premier
International Research (WPI) Center Initiative on Materials
Nanoarchitronics (MANA), MEXT, Japan, the National High
Technology Research and Development Program of China (863
program: 2009AA03Z309) and the WCU (World Class University) program through the National Research Foundation
(NRF) of Korea funded by the Ministry of Education, Science
and Technology (R31-10013).
This journal is The Royal Society of Chemistry 2012

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