Recent Developments in Molecularly Imprinted Nanoparticles by Surface Imprinting Techniques
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Transcript of Recent Developments in Molecularly Imprinted Nanoparticles by Surface Imprinting Techniques
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Review
Recent Developments in MolecularlyImprinted Nanoparticles by SurfaceImprinting Techniques
Xiaochu Ding,* Patricia A. Heiden*
Molecularly imprinted polymer nanoparticles (MIPNPs) are an increasingly important area ofresearch with potential in applications such as biosensors, solid phase extractions andbioassays. Advantages over the traditionalmolecularly imprinted polymers typically include ahigher binding capacity, greater selectivity and affinity for target species, and aqueouscompatibility. Recent research efforts have sought to impart MIPNPs with additionalcapabilities by introducing nanoparticle size-control, stimuli-responsiveness, biocompatibili-ty, and optoelectronic properties. This short review describes the molecular imprinting
principle and then discusses recent advances inthe field of MIPNPs with particular focus onsurface polymerization techniques to imprintboth small and macro molecules.X. Ding, P. A. HeidenDepartment of Chemistry, Michigan Technological University,Houghton, MI 49931, USAE-mail: [email protected]; [email protected]
� 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim wileyonlinelibrary.c
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1. Introduction
Natural receptors associated with enzymes, antibodies,
active proteins, etc. all possess excellent binding selectivity
and affinity and are widely studied so their properties
can be reproduced for use in chemo/biosensors,[1–3] bio-
assays,[4–7] and biomedical diagnostics.[8,9] However,
natural receptors also have significant disadvantages that
limit their commercial use. These disadvantages include
poor physical/mechanical stability and high cost to
produce, and their use is limited to moderate conditions
to preserve their binding specificity.[10] In the past few
decades, chemists and biologists have invested time and
effort to develop artificial receptors that possess similar
specific binding and affinity properties for targetmolecules
but are easy to produce and are cost-effective. Molecular
imprinting is an efficient strategy to achieve these
objectives.
Molecularly imprinted polymers (MIPs) are designed to
possess antibody-like selectivity similar to their natural
analogs, but unlike those materials, can be designed to
possess physical/mechanical stability, resistance to harsh
conditions (e.g., high temperature, pressure, acids, bases,
and some organic solvents), and be produced more easily
and economically.[11–13] The key to an effective MIPs is to
produce a ‘‘device’’ that possesses recognition sites with
appropriate spatial arrangement and specific affinity for
a template. The standard approach to prepare MIPs is
illustrated schematically in Figure 1. [11]
Figure 1 shows how vinylic monomers, mixed together
with a ‘‘template,’’ produce a pre-polymerization complex.
The monomers are then polymerized around the template,
together with a difunctional cross-linker, to form a highly
cross-linked polymer network that holds the template
shape. The template is then removed by extraction or
chemical cleavage to leave the shape-specific binding sites.
MIP materials prepared by traditional methodologies
such as bulk polymerization, precipitation polymerization,
and emulsion polymerization, have been extensively
studied for use in solid-phase extractions,[14] chemo/
biosensors,[15,16] andanalytical chemistry.[17,18] Thedimen-
sionsofMIPmaterialsprepared in thesewaysusually range
from ‘‘bulk’’ to micron sized particles. For bulk MIPs, after
the imprinting process is completed, they are usually
ground into a powder and ‘‘sized’’ by passing them through
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Xiaochu Ding joined the Department of Chemistryat Michigan Technological University in Au-gust 2008. He studied in the research group ofProf. Patricia A. Heiden, andwill receive his Ph.D. inAugust 2013. He received his B.Sc. in Chemistry(SouthwestUniversity forNationalities, China) andhisM.Sc. in Chemistry (ZhejiangUniversity, China).His research interests include design and synthesisof functionalized organic and inorganic nano-particles for smart self-assembly and self-assem-bled nanofibers for tissue engineering, targetedand controlled drug delivery and biosensing.
Professor Heiden received her B.Sc. in Chemistry(Wright State University, Dayton, Ohio, USA) andher Ph.D. in Polymer Science at University of Akron(Akron,Ohio,USA)working in the researchgroupofDr. Frank Harris. She joined the Department ofChemistry at Michigan Technological University in1994 where she is now a Professor of Chemistryresearching nanoparticles, nanofibers, and bio-polymer use and modification for sustainablematerials.
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X. Ding, P. A. Heiden
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filters having the pre-determined mesh size to isolate the
particles in thedesired size range.Approximatedimensions
of these particles are typically �10�2–10�6m, but other
sizes arepossible. Theparticles typicallypossessa relatively
small number of binding sites on or near the surface
because of the small ratio of surface area to volume.[13] For
perspective, the surface area to volume ratio (S/V ratio)
increases ten times as particle size decreases from 1mm to
100nm.
Although traditional bulk ormicro-sizedMIPs are easy to
prepare, several factors, arising from their highly cross-
linked structures and irregular shapes, limit their applica-
tion as artificial receptors. The specific disadvantages
of bulk and micro-sized MIPs include: (1) difficulty of
removing target molecules from interior binding sites;
(2) the rebinding capacity is limited by the small number of
binding sites on/near the surface; and (3) target molecules
are easily hindered from accessing binding sites deep in
the interior of the particles.[19] These problems need to
be resolved before a truly successful MIP system can
be introduced for recognition of macromolecules (e.g.,
proteins).
Molecular recognition systems for proteins are more
complex than those for small molecular recognition
systems. This is because several other essential factors
must be taken into consideration when preparing MIPs for
protein recognition. First, the reaction conditions must be
carefully controlled so as to make the structure of the
template close to that of the rebinding protein as it exists
in the biological environment where it is to be used,
because themorphology of the protein can be significantly
affected by temperature, pH, ion concentration, etc.[20,21]
Second, the slow rebinding kinetics of a target protein to
binding sites that are deep in the MIP interior, must be
overcome. Molecular imprinting on the surface of particles
is the preferred method to accomplish this. This is done, by
imprintingatemplateproteinonto thesurfaceofmicro-size
particles or solid substrate surfaces.[22–25] The process is
illustrated in Figure 2. First, a solid substrate or particle, a
protein complex (i.e., a protein that is already complexed to
vinylicmonomers that bear functional groups that are able
to interact with the protein), and a difunctional crosslinker
are combined. Then, the copolymerization and cross-
linking are initiated. The resulting product now contains
Figure 1. A schematic representation of a molecular imprinting proc
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the solid support, or ‘‘substrate module,’’ with the
complexed proteins and the cross-linked shell. The com-
plexed protein is then removed, leaving the surface binding
sites that can rebind to the target proteins.
This technique successfully places binding sites on
substrate surfaces after removing the template, but the
density of surface binding sites is still limited due to the
small surface area to volume ratio of these conventional
MIPs.
Nanostructured materials (e.g., nanoparticles) are now
among the most researched alternatives to overcome the
drawbacks associated with conventional MIPs. Indeed, a
search of molecularly imprinted polymer nanoparticles
(MIPNPs) throughWebof Science shows329publications in
last decade. However, almost half of these (158 publica-
tions) are issued in the last twoyears. BecauseMIPNPshave
such a high surface-to-volume ratio the imprinted binding
sites are necessarily created on/near the nano-material’s
surface. This improves the rebinding capacity and imparts
faster rebindingkinetics for the targetmacromolecules, and
so is an efficient strategy to overcome the limitations of
bulk MIPs.
ess. Adapted with permission.[11] Copyright 2004, Springer.
: 10.1002/mame.201300160
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Figure 2. A surface molecular imprinting process to giveprotein recognition sites on micro-size particle surfaces orflat substrate surfaces is shown. The process if followed byremoval of the template proteins to leave recognition sites onthe substrate surface.
Recent Developments in Molecularly Imprinted Nanoparticles . . .
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Several recent review papers have described strategies
for the preparation of molecularly imprinted polymer
micro and nanoparticles.[26–31] The advantages of MIPNPs,
compared with traditional MIP materials, have also been
widely discussed with respect to bioassay, biosensor,
environmental analysis, and biomimetic catalytic applica-
tions.[32–37] This review differs from these prior reviews in
its focus on the recent advances inMIPNPpreparation,with
particular attention paid to surface polymerizations on
various nanoparticle (NP) substrates to enhance small
molecule andmacromolecular imprinting and recognition.
These recently developed techniques effectively create
the recognition sites on MIPNP surfaces so that they
are accessible. Also, by selection of an appropriate NP
substrate additional capabilities can be given to the
MIPNP systems, such as magnetic responsiveness for ease
of separation or optoelectronic properties for detection.
Table 1 lists the most commonly used NP substrates,
their surface functional groups, and the corresponding
MIPNP preparation techniques that we are going to
discuss in this review.
The first half of this review focuses on imprinting
and recognition processes, and applications for species
imprinted with small molecules. The second half of
the review addresses imprinting and recognition, appli-
cations, and the latest advances in macromolecular
imprinting and recognition, including discussing the
specific perceived advantages of MIPNPs compared to
conventional MIPs. To avoid overlap within this review,
some of the new advances of MIPNPs described with
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the small molecularly imprinted MIPNPs will not be
addressed a second time with examples of macro-
molecular imprinted systems.
2. Recent Advances in MIPNPs for SmallMolecule Recognition and TheirApplications
The traditional polymerization strategies used to obtain
MIPNPs that are rich with surface binding sites include
precipitation, emulsion, mini-emulsion, and surface-
initiation on polymer seeds.[38] In recent years more
versatile techniques have been proposed and tested for
thepolymerization of a surface layer on solidNP substrates.
These newmethods give excellent control over MIPNP size
by selection of suitable NP substrates and controlling the
thickness of the imprinted layer. These methods can also
allow use of stimulus-response and biocompatible MIPNP
shell materials for different applications.
2.1. MIPNPs on Silica NP Substrates
Silica NPs are popular substrates for surface imprinted
MIPNPs in part because the synthesis of silica NPs
themselves is a well-established process. Many simple
synthetic methods are described in the literature that give
the conditions that allow the MIPNP particle diameter to
be controlled anywhere within a range of micrometers
to nanometers. Also, silica possesses numerous hydroxyl
groups on its surface, and these are readily employed as
grafting sites for the surface polymer. Therefore, it is a
relatively straightforward matter to prepare silica
MIPNPs with a controlled size and surface.[39,40] There is
also a wide range of reactions and monomers that can
be used for the surface functionalization of silica NPs.
Finally, the chemical/mechanical stability, non-toxicity
and biocompatibility of silica all make it an attractive
substrate for use in many fields.[41]
Silica NPs are probably most often functionalized with
organic or inorganic vinyl functional groups,[42–46] iso-
cyanates,[47] and amine groups.[48,49] The next step is the
imprinting process, using an existing template followed
by polymerization with functional monomers and then
cross-linkers. These methods yield a high density of
effective binding sites on the silica NP surface with good
accessibility, selectivity, and a high rebinding efficiency for
the target molecules.[50,51]
Several variations on the surface imprinting technique
have been described in the recent literature. For example,
Guan et al.[52] reported imprinting 2,4,6-trinitrotoluene
(TNT) molecules on the surface of amine-functionalized
silica NPs (130nm) via a layer-by-layer technique. In this
method, instead of copolymerization of vinylic monomers,
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Table 1. Summaries of nanoparticle substrates, surface functional groups, and MIPNP preparation techniques.
Substrate
Surface functional
group
MIPNP preparation
technique Refs.
Silica NP Vinyl Surface polymerization [42,43,45,46,104]
OH Sol–gel [44]
Isocyanate Condensation polymerization [47]
Amine Layer-by-layer [48,49,52]
RAFT RAFT polymerization [22]
Template molecule Pickering emulsion polymerization [53]
Au NP Thioaniline Electropolymerization [56–58]
Magnetic NP ATRP ATRP polymerization [64,96,97]
Vinyl Radical polymerization [65–67,94]
Amine Sol–gel [98,99]
None Self-polymerization of DPA [92]
None Phase inversion method [103]
QD Vinyl Radical polymerization [73,107]
Amine Cross-linking reaction, sol–gel [74,76]
OH Sol–gel [77,78,82]
Carboxylic acid Sol–gel [79]
ATRP ATRP polymerization [80]
None Encapsulation [81]
Template molecule Sol–gel [108]
None Phase inversion [109]
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X. Ding, P. A. Heiden
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a glutaraldehyde (GA) mediated covalent assembly of
gelatin was used to form the imprinted layers. The final
MIPNPs possessed a diameter of 180nm. The authors
found that the rebinding capacity of TNT changed
nonlinearly with the layer number of the TNT-imprinted
gelatin layers, but observed that three layers of
imprinted gelatin yielded the highest rebinding capacity
in this study. The significanceof this layer-by-layer surface
imprinting technique is that it expands the range of
materials that can be used to form the surface-imprinting
layer from vinylic monomers to biopolymers (e.g.,
chitosan, proteins, enzymes), and many of these can be
used in aqueous media by a simple covalent assembly
between GA and the amine groups of those biopolymers.
This technique also allowed excellent control of the
thickness of the imprinted layers by changing the number
of applied layers.
Another relatively simple procedure was employed
by Zhu et al.[44] to imprint bisphenol A (BPA) on a silica
NP surface using a double-sol–gel method. In the first
step tetraethylorthosilicate (TEOS) was hydrolyzedwith
ammonium hydroxide to obtain spherical silica NPs
with a size of 400 nm. Then the NPs were washed with
anhydrous ethanol. In the second step, the imprinting
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process was performed by dispersing the silica NPs
in methanol with the aid of sonication. Then the
template molecules (BPA), the functional monomers
3-aminopropyltriethoxysilane (APTES) and TEOS, and
a small amount of acetic acid were combined. This
mixture was stirred and polymerized at room tempera-
ture for 18 h. After repeatedly washing the product
with a mixture of methanol and 6 M HCl (1:1 v/v) to
remove the templates from MIPNPs, the researchers
tested the efficacy of these MIPNPS using solid phase
extraction, which showed these BPA-imprinted silica
MIPNPs possessed high absorption capacity, high
selectivity, and fast rebinding kinetics for the target
BPA molecules.
Chang et al.[22] recently demonstrated an attractive
method to imprint 2,4-dichlorophenol (2,4-DCP) on silica
micro beads having a polymer layer thickness of�2.27nm.
The author coupled an alkyne-terminated reversible
addition-fragmentation chain transfer agent (RAFT) to
azide-functionalized silica beads via a click conjugation.
The author then grafted a thin, uniform nanofilm on the
silica NP surface that was imprinted with 2,4-DCP. This
technique allowed effective control of the thickness of
the imprinted layer because it used RAFT, a controlled
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Recent Developments in Molecularly Imprinted Nanoparticles . . .
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radical polymerization, and demonstrated high selectivity
and fast rebinding kinetics towards the target molecule,
2,4-DCP.
Another approach, called interfacial imprinting, was
recently reported by Shen et al.[53] This process, illustrated
inFigure3, employedaPickering emulsionpolymerization
method. The template molecules are first immobilized on
the surface of silica NPs (Figure 3a), often prepared from
TEOS, and these NPs are used to stabilize monomer
droplets (80–240mm diameter) to form a stable oil-in-
water emulsion (left side of Figure 3b). The monomer
droplets, composed of methacrylic acid (MAA) and
ethylene glycol dimethacrylate (EGDMA), are then poly-
merized and cross-linked with azobis(isobutyronitrile;
AIBN; Step 1 in Figure 3b). The silica NPs on the surface of
the microsphere were then removed by stirring the
particles in a solution of HF (30wt%) for 12 h (Step 2 in
Figure 3b), leaving the imprinted pores with specific
affinity for the target molecules. This new type of MIP
microsphere possesses a well-controlled hierarchical
structure with large pores for easily accessible binding
sites. Use of the silica-bound templatemolecule-I (left side
of Figure 3a) allows for selective recognition of a series
of analogous compounds that have the isopropylamino-
propanediol epitope as a common motif, e.g., atenolol,
metoprolol, pindolol, and propranolol, while use of the
silica bound template molecule-II (on the right side of
Figure 3a) gives non-specific recognition to the same series
of compounds. MIP microspheres prepared in this way
Figure 3. Illustration of an interfacial molecular imprinting on silicaimmobilized on silica NPs, while b) showsmonomer droplets in water,These are imprinted by copolymerizing with cross-linker in Step 1, to fophase is removed by stirring in an HF solution (30wt%). The interior of(b). Figure 3a adapted with permission.[53] Copyright 2011, American
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possess hydrophilic surfaces, enabling them to be used in
aqueous media.
2.2. MIPNPs on Au NP Substrates
Gold nanoparticles (Au NPs) are another popular substrate
on which to assemble an imprinted layer, because Au NPs
possess a surface plasmon resonance (SPR) which is
sensitive to interparticle distance, and to the absorption
of some target molecules or biomolecules that change
the thickness of the surface layer.[54,55] The SPR feature
makes Au NPs an attractive substrate for use in the field
of biosensors and biodiagnostics.
The value of the SPR is illustrated in recent publications
by Willner’s group.[56–58] They demonstrated the ultra-
sensitive SPR detection of hexahydro-1,3,5-trinitro-1,3,5-
triazine (RDX). The multi-step process, illustrated in
Figure 4,[56] begins with the electropolymerization of
thioaniline-functionalized Au NPs (3.5 nm) onto an Au
electrode that is already coated with a thioaniline-
monolayer. The electropolymerization is done in the
presence of an imprinting template molecule (not shown
in Figure 4a), to give a cross-linked bisaniline network
bound to Au NP substrates that are themselves now
bound onto the Au electrode surface. Because RDX
possesses low solubility in the aqueous electropolymeriza-
tion solution, the authors did not use RDX itself as the
template, but instead used Kemp’s acid as an analogous
template molecule for RDX. Figure 4b shows the extraction
NP-stabilized emulsion drops; a) shows template molecules (I or II)stabilized by the silica NPs with the immobilized templatemolecules.rm cross-linked polymeric microspheres, and then in Step 2, the silicaone of the imprinted sites is expanded and shown on the right side ofChemical Society.
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Figure 4. A schematic illustration that shows MIPNPs with ultrasensitive SPR detection of RDX. a) Thioaniline and mercaptoethane sulfonicacid functionalized Au NPs (3.5 nm) are electropolymerized onto a thioaniline-monolayer-modified Au electrode to form ‘‘sponge’’composites of bisaniline-cross-linked Au NPs associated with Au surface. b) Kemp’s acid-imprinted MIP Au NP composites forrecognition sites for RDX analysis.[56]
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X. Ding, P. A. Heiden
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of Kemp’s acid leaving highly sensitive receptor sites for
RDX.
Theauthors foundthatKemp’sacidwasasuitableanalog
for RDX that gave highly sensitive detection of RDX.
This was because their research showed that RDX bonds to
receptor sites through p-donor–acceptor interactions
between RDX and the bisaniline units.[56] Their Au-MIPNPs
generated measurable reflectance changes in the SPR
spectrum with a detection limit as low as 12 fM, which
is 4� 105-fold lower than a non-imprinted sensing matrix.
In subsequent work they also demonstrated chiroselec-
tivity through imprinting of L-glutamic acids or D-glutamic
acids by electropolymerization of thioaniline and cysteine
functionalized Au NPs, using bisaniline as a bridge to form
the chiroselective recognition sites.[57] The chiroselectivity
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results from zwitterionic interactions and hydrogen
bonding between the amino acids and the cysteine units.
The rebinding of target chiral amino acids could be
distinguished by the SPR spectrum with a detection limit
of 2 nM. Their studies also showed Au NPs with bisaniline
bridges in themolecularly imprinted composites displayed
quasi-reversible redox properties. When at E< 0.12V,
versus Ag as a quasi-reference electrode (QRE), the bridges
existed in the reduced bisaniline (p-donor state). However,
at E> 0.12V versus Ag QRE, the bridges changed to
the quinoid (p-acceptor state).[58] This potential-induced
reversible uptake and release of p-acceptor molecules
suggests potential applications in chromatographic sepa-
rations, controlled release systems, and removal of
pollutants.
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Recent Developments in Molecularly Imprinted Nanoparticles . . .
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As previously stated, Au NPs possess an SPR which is
highly sensitive to changes in the environment around
the particles. So this feature, along with the chemical
stability, ease of surface modification, and wide range of
controllable particle size at which Au NPs can be synthe-
sized, make Au-MIPNPs and their nanocomposites attrac-
tive substrates for use as sensors for molecular detection
inpreference toothermetallicNPs.[59] A recentmini-review
described the strategies and advantages of Au-MIPNP
composite films used as sensors for various applications
and detections.[60]
2.3. MIPNPs on Magnetic NP Substrates
Magnetic NPs are also of interest as substrates for the
preparation of small molecularly imprinted MIPNPs that
combine the ability to recognize target molecules with
magnetic responsiveness. In the last decade, 113 publica-
tions describe magnetic MIPNPs, and more than half of
thesedescribe smallmolecularly imprintedmagnetic Fe3O4
NPs for uses like separation, solid phase extraction, and
sensors, as described in several recent reviews.[32,61–63] The
strategies used to prepare magnetic NPs that possess
surface functionalities (e.g., vinyl groups, amine groups, or
groups able to function as RAFT chain transfer agents) are
described in the section on macromolecular imprinted
magneticNPs. This is donebecauseof their value inprotein-
imprinted MIPNPs. In this section the focus is on
representative methods to imprint the NP surfaces and
applications of the imprinted particles.
A ‘‘grafting from’’ technique is frequently used to
fabricate molecularly imprinted layers on magnetic NP
surfaces. Generally, the magnetic NP surface is first
decorated with a silica layer, because this provides an
abundance of functional groups that, as described previ-
ously, can be easily converted to other desirable groups.
Figure 5. A representative surface imprinting process is shown that uses a ‘‘graftingfrom’’ technique on ATRP-functionalized magnetic NPs.
Then the magnetic NPs can be further
functionalized with species able to serve
as a radical chain transfer agent for RAFT,
or atom transfer radical polymerization
(ATRP). This step is followed by a surface
imprinting process to create specific
binding sites. For example, Liu et al.[64]
reported a procedure to make Fe3O4 NP
surfaces suitable for ATRP. The process
began by reaction of Fe3O4 NP surfaces
with TEOS, in thepresence of ammonium
hydroxide, to obtain Fe3O4@SiO2 NPs.
Thiswas followed by additional hydroly-
sis with ammonium hydroxide in
the presence APTES to form Fe3O4@-
SiO2@NH2 NPs. Finally, the ATRP initia-
tor, 2-bromoisobutyryl bromide, was
immobilized on the Fe3O4@SiO2@NH2
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NP surfaces via reaction of the acyl bromide with the
surface amine groups. Then suitable monomers were
polymerized from the surface, in thepresence of the desired
template, in this case the antibiotic pefloxacin mesylate
(PEF-M), to yield the imprinted magnetic MIPNPs, having a
particle size of 500nm and an imprinted layer having a
depth of 18nm. Testing of PEF-M-imprinted Fe3O4 NPs
showed significant specific affinity towards the target PEF-
M in aqueous media and high rebinding capacity. The PEF-
M-imprinted Fe3O4 NPs were highly magnetic (41.4 emu
g�1) and responded to external magnetic fields allowing
rapid separation of the MIPNPs after bonding with PEF-M
molecules from an egg sample. The general ‘‘grafting
from’’ technique on ATRP-functionalized magnetic NP
surface is shown in Figure 5.
‘‘Grafting to’’ techniques are also employed with
Fe3O4 NPs coated with a SiO2 shell obtained by sol–gel
methods, using TEOS and 3-methacryloxypropyltri-
methoxysilane (MPS), to form Fe3O4@SiO2 NPs with
vinyl groups on the surface of the NPs. Then the
molecularly imprinted layer is formed on the surface
by copolymerizing the vinyl groups with functional
monomers and cross-linkers in the presence of the
desired templatemolecules. This approachwas employed
in several recent publications.[65–67] For instance,
Kong et al.[67] showed that sulfamethazine molecules
were effectively imprinted on vinyl-functionalized
Fe3O4@SiO2 NP surfaces using MAA as the functional
monomer, EGDMA as the cross-linker, and AIBN as
the initiator. The resulting MIPNPs had high binding
capacity and fast re-binding kinetics.
A schematic representation of this ‘‘grafting to’’ tech-
nique on the vinyl-functionalized NP surface will be
illustrated in the macromolecular imprinting section.
Additional surface functionalization methods and newer
surface imprinting technologies using magnetic NPs will
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also be discussed in the section on macromolecular
imprinted MIPNPs.
2.4. MIPNPs on Quantum Dot Substrates
Quantum dots (QDs), such as CdS, CdTe, CdSe, ZnS, etc., are
often used as biolabels or to fabricate chemo/biosensors
because of their optoelectronic characteristics.[68–71] The
techniques to fabricate QD-based MIP chemo/biosensors
were developed only recently. The recognition cavities
capture templates that quench the photoluminescence
emissions from the QDs due to fluorescence resonance
energy transfer between QDs and template molecules.[72]
The QD-based/MIP devices can usually be classified into
four categories for small molecule detections.
The first, and most traditional, approach is to embed
QDs into MIP matrices during the molecular imprinting
process. This is done by copolymerization using vinyl
functionalized QDs along with the desired templates,
functional monomers, and cross-linkers. For example,
QD embedded MIPs were synthesized using MAA as
the functional monomer, together with 4-vinylpyridine
functionalized CdSe–ZnS core-shell QDs, and EGDMA as a
cross-linker, in the presence of caffeine as the template
molecule.[73] A post-treatment method can also be used
to embed surface functionalized QDs on MIP matrices
by cross-linking amine-functionalized QDs with MIP
matrices that contain carboxylic acid groups.[74] This
traditional sort of QD embedded MIP material can be cast
into films or ground into particles for molecular detection,
as desired for the intended application. The general process
Figure 6. The traditional QD-embeddedMIPs. a) Molecular imprintingtemplate and cross-linker; b) post-treatment using an amine-functio
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to prepare these two QD-embedded MIPs is illustrated in
Figure 6.
The second approach uses a direct surface imprinting
technique on the QD substrate to form an imprinted outer
layer on the QD-MIPNPs. The binding cavities on the outer
layers can be generated by a surface polymerization
technique if vinyl-functionalized QDs are used,[75] or by
sol–gel methods if the QD surfaces possess amines,[76]
silanols,[77,78] or carboxylic acid groups.[79] However,
because of the small size of the QDs, some of the imprinted
outer layers will actually be coated on QD aggregates
comprised of several QDs.[75,76] The surface imprinting
process on vinyl-functionalized QD is similar with other
vinyl-functionalized NP substrates, except QDs slightly
forming aggregations.
In the third category the MIP layer is formed on QD-
embedded nanoparticle substrates to fabricate multi-
functional biosensors with the following structure:
QDs@NPsubstrate@MIP. For example, ZnO nanorods can
be surface functionalized with an ATRP initiator (2-
bromoisobutyryl bromide) and then surface polymeriza-
tion canbe carried out in presence ofmagneticNPs (g-Fe2O3
NP), functional monomers (MAA), cross-linkers (EGDMA),
and templates to form a ZnO@Fe2O3@MIP biosensor that
can give selective antibiotic detection.[80] This is a new
type of biosensor that possesses functionalities that
are reported to be able to give selective recognition,
separation, and detection. Although these biosensors
have not yet been well-studied, they reportedly meet
market requirements for a device suitable for high
throughput analysis, such as might be used in food supply
polymerization using a vinyl-functionalized QD, a vinylic monomer, analized QD, a polymer with carboxylic acid groups and a template.
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Figure 7. General procedure for the preparation of multi-functional QD-embedded magnetic MIPNPs.
Recent Developments in Molecularly Imprinted Nanoparticles . . .
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management, bio-diagnostics and environmental protec-
tion. The general procedure of preparing QD-embedded
magnetic MIPNPs is shown in Figure 7.
The final category of QD-based MIPs consists of QD-
embeddedMIP nanocomposites. Either surfacemodified or
non-modified QDs can be used to fabricate this type of QD-
basedMIP sensor. This approachwas used by Zhao et al.,[81]
who used an ultrasonication-assisted encapsulation
method that yielded Mn–ZnS embedded QD-MIP nano-
composites for diazinondetection. Thiswas accomplishedby
dispersing non-functionalized Mn–ZnS QDs, the template
(diazinon), and a pre-synthesized copolymer (poly(styrene-
co-methacrylic acid)) into chloroform to give a transparent
solution that was then injected into deionized water under
ultrasonication andwithmagnetic stirring for 6min to form
a diazinon-imprinted QD-MIP nanocomposite suspension.
After washing and collecting by centrifugation, this easily
prepared QD-MIP nanocomposite was tested against diazi-
non in water and showed a good linear correlation, in a
concentration range of 50–600ngmL�1, with a sensitive
detection limit (down to 50ngmL�1 of diazinon in water),
and with excellent selectivity to the analyte.
Another example of this approach used lambda-cyhalo-
thrin (LC) as the template molecule and employed
functionalized QDs. The QD-MIP nanocomposite was
fabricated using APTES functionalized CdSe QDs that can
copolymerize with TEOS in the presence of the template
molecules.[82] The decision to use functionalized or non-
functionalized QDs depends on the type of template
molecule to be imprinted. Surface functionalized QDs are
preferred as assistant monomers if their interaction with
template molecules can increase imprinting efficiency and
rebinding selectivity.
2.5. MIPNPs Prepared by Modified Precipitation
Polymerization and Mini-Emulsion Polymerization
Techniques
Traditional polymerization techniques, such as precipita-
tion polymerization and emulsion/mini-emulsion poly-
merization, have also been useful for the preparation of
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MIPNPs for small molecular imprinting, though they have
beenperformedwithmodifications on the traditional route
so they yield MIPNPs. Yang et al.[83] designed a variation
of precipitation polymerization, called distillation precipi-
tation polymerization, to give propranolol-imprinted
MIPNPs within 3h, in contrast to the 24h reaction time
that is common in traditional precipitationpolymerization.
The authors also synthesized core–shell structuredMIPNPs
by this method, where the core contained imprinted
binding sites while a hydrophilic shell was expected
to prevent non-specific adsorption of biomolecules (i.e.,
protein) and allow the small target molecules to enter the
binding sites in the hydrophobic core. Such MIPNPs are
potentially useful for extraction of small organicmolecules
from complex biological samples.
Pan et al.[84] used surface-initiated RAFT polymerization
to introduce poly (N-isopropyl acrylamide; PNIPAAm)
brushes onto MIPNP surface. This not only improved the
surface hydrophilicity, that helped stabilize the imprinted
NPs, but also imparted stimuli-responsive properties to the
MIPNPs’ surface layers.
Recently, Esfandynari-Manesh et al.[85] showed that
carbamazepine-imprinted MIPNPs, prepared by mini-
emulsion polymerization, could be utilized as a drug
delivery system giving a sustainable release of carbamaze-
pine with a higher binding level and slower release rate
than non-imprinted NPs. The study showed that the ratio
of template to functional monomer was a key factor
to obtain the best drug affinity. The release rate of the
loaded drug lasted for more than eight days in 1wt%
sodium dodecyl sulfate aqueous solution.
3. Recent Advances in MIPNPs for ProteinRecognition and Their Applications
Despite the success of small molecular imprinting techni-
ques and their applications in various fields, macromolec-
ularly imprinted MIPNPs remains challenging because of
the difficulties associated with conformational stability,
mass transfer hindrance, and slow rebinding kinetics.
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In recent years though, significant progress has been
reported in macromolecular imprinting technologies,
especially for applications in bioseparations, biosensors,
and biodiagnostics.
While surface imprinting is an efficient technique to
create binding cavities for proteins on the outer layer of
MIPNPswith good accessibility and fast rebinding kinetics,
the conditions used to accomplish the imprinting process
must be carefully considered. The conformational stability
of proteins is sensitive to their surrounding conditions,
including temperature, pH, ion concentration, surfactant
and its concentration, the ratio of template to functional
monomers, the types of functional monomers used, and
initiator type.[55,86–88] Identifying those reaction conditions
that ensure the conformational integrity of a template
protein and increase binding sites on or near the MIPNP
surface, and that ensure good accessibility, are crucial for
any useful protein recognition system. Several review
papers have described suitable conditions for synthesizing
protein-imprinted MIPNPs.[86,89,90]
A general procedure for protein imprinting on vinyl-
functionalized NP surfaces is shown in Figure 8. The
substrate canbe silicaNPs,magneticNPs, orQDsdepending
on the intended use. Functional monomers, such as MAA,
acrylamide (AAm), and N-isopropylacrylamide (NIPAAm),
are usually used so they can form electrostatic, hydrogen-
bonding, and hydrophobic interactions with appropriate
domains of the template protein. EGDMA or bisacrylamide
(BIS) are often used as cross-linkers to copolymerize with
the functional monomers to give the cross-linked protein-
imprinted layers on theNP substrate surface. The process is
conducted in aqueous media and under appropriate
conditions of pH, salt concentration, reaction temperature,
and with an appropriate ratio of functional monomers to
protein templates.
The sections below describe recent advances in protein
imprinting according to different types of substrate
Figure 8. A general procedure to produce a protein-imprintedMIPNP on a vinyl functionalized NP substrate surface by a surfaceimprinting technique.
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categories, and improvements in newer preparation
methods will also be discussed.
3.1. Protein-Imprinted MIPNPs on Magnetic NP
Substrates
Magnetic NPs such as Fe3O4 are among the most useful
substrates for protein imprinting because the magnetic
responsiveness allows these MIPNPs to be easily separated
from complex samples simply by applying an external
magnetic field. This avoids the tedious separations involv-
ing centrifugation or filtration. These NPs are often
prepared by a co-precipitation[91] or a solvothermal[92]
reaction route, usually yielding NPswith a diameter below
25nm that possess superparamagnetism. The major
applications for these MIPNPs are separation and
detection.[93]
One of the most common methods to prepare protein-
imprinted magnetic MIPNPs employs vinyl-functionalized
magneticNP substrates. For example, Jing et al.[94] obtained
a 20nm thick lysozyme-imprinted layer on vinyl function-
alizedFe3O4@SiO2NPswithatotaldiameterof172� 28nm
and with a rebinding capacity of 0.11mg lysozyme/mg
magnetic MIPNP for the target proteins. These researchers
prepared the Fe3O4NPs using a co-precipitation route. First,
they dissolved FeCl3 � 6H2O and FeCl2 � 4H2O in deionized
water with continuous mechanical stirring under a
nitrogen atmosphere. They obtained the desired NPs after
heating to 80 8C with a dropwise addition of ammonium
hydroxide, and maintaining the reaction at that tempera-
ture for 30min. The as-made Fe3O4 NPs were further
modified with a thin layer of silica having silanol surface
functionality via a sol–gel reaction of TEOS in an ammoni-
um hydroxide solution. Then the Fe3O4@SiO2 NPs were
functionalized with vinyl groups obtained by the hydroly-
sis of MPS in acetic acid solution.[94,95] In the subsequent
imprinting process, the selection of the functional mono-
mers and cross-linker must take into consideration the
specific template protein to be used. Although no specific
monomersaredesignated foragivenprotein, combinations
of hydrophobic and hydrophilic monomers are typically
selected that are capable of forming hydrophobic, ionic, or
hydrogen bonding interactions to suitable domains of
the protein. In this study, the authors opted to use MAA
and AAm as functional monomers to form electrostatic
and hydrogen bonding interactions with lysozyme as
a template protein, and N,N0-methylenebisacrylamide
(MBAAm) was employed as a cross-linker. The copolymeri-
zationwas performed in phosphate buffered solution (PBS)
to give the imprinted layer, and then the template protein
waswashed awayby 1MNaCl and deionizedwater to leave
the specific binding sites for lysozyme.
Magnetic NPs have also been prepared with suitable
functionalities so that the imprinted layers canbe prepared
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Recent Developments in Molecularly Imprinted Nanoparticles . . .
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under aqueous conditions by ATRP polymerization.[96,97]
For example, Gai et al.[96] prepared a lysozyme-imprinted
layer with a thickness of 15nm on ATRP-functionalized
Fe3O4@SiO2NPswitha total diameter of 120nm.Again, the
Fe3O4 NPs were surface-modified with a silica–NH2 layer,
introduced by hydrolysis in an APTES/ethanol/water
solution (12 h at 40 8C) to give Fe3O4@SiO2–NH2 NPs. The
Fe3O4@SiO2–NH2 NPs were then reacted with 2-bromoi-
sobutyryl bromide to anchor the ATRP active agent, using a
mixture of tetrahydrofuran (THF) and triethylamine (TEA)
for 20min in an ice bathunder nitrogenatmosphere, giving
Fe3O4@SiO2–ATRP. Finally, the Fe3O4@SiO2–ATRP was
used as the initiator, and combined with the functional
monomers (NIPAAm and AAm) and cross-linker (MBAAm),
along with the template proteins (lysozyme), in a typical
ATRP polymerization process, to give the surface
imprinted layer. This lysozyme-imprinting process was
performed on the magnetic NPs in PBS solution at room
temperature, followed by removal of the lysozyme
template using acetic acid (10% v/v)–SDS (10%w/v), to
give lysozyme-imprinted MIPNPs having excellent desorp-
tion and extraction capability. This study demonstrated
that ATRP-mediated surface imprinting techniques could
be performed under mild reaction conditions and in
aqueous media, and is important because these methods
allow better control over the thickness of the imprinted
layers than non-controlled radical polymerization meth-
ods. This is advantageous because the recognition sites of
thinner imprinted layers are more easily accessible for
target proteins.
Figure 9. Surface imprinting of BHb on amine-functionalized Fe3O4 NPprocess.
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Amine functionalized Fe3O4 NPs can also be designed to
covalently bond to a template protein, such as bovine
hemoglobin (BHb).[98,99] This is done using GA that bonds
to the BHb forming an imine bond. Then a simple sol–gel
process, using a mixture of TEOS and APTES, is followed
to complete the imprinting process. By this method, Kan
et al.[98] reported that the thickness of BHb-imprinted layer
was about10nm,whichwas close to the spatial size of BHb,
indicating that the imprinted sites were located near the
surface of the magnetic MIPNPs. The imprinted BHb was
then extracted using a mixture of deionized water and
methanol (1:3.2 v/v)with sodiumbicarbonate (�3.0mmol)
at 25 8C for 20h with mechanical stirring under nitrogen
atmosphere. The protein adsorption tests showed a rapid
rebinding equilibrium, achieved within 1h, with a rebind-
ing capacity of 10.52mg BHb g�1magneticMIPNP. Figure 9
shows the imprinting process of BHb on the surface of
amine-functionalized Fe3O4 NPs, as described above.
Recently, 3-aminophenylboronic acid (APBA), and dop-
amine (DA) have been explored as new functional
monomers for use in magnetic MIPNPs.[92,100,101] DA
is a neurotransmitter that is biocompatible and bio-
degradable. DA contains several functional groups and
can self-polymerize (to PDA) in a weakly basic aqueous
solution, as shown in Figure 10. [102] Zhou et al.[92] used DA
as the functional monomer to imprint human hemoglobin
to the surface of a magnetic NP by mixing the Fe3O4 NPs
(100mg) and hemoglobin (20mg) in 20mL Tris buffer
(10mM, pH 8.0). After mechanically stirring for 2h at room
temperature, DA (40mg) was added and the reaction was
surface based on covalent bond of glutaraldehyde-amine and sol–gel
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Figure 10. Illustration of dopamine self-polymerization in weaklybasic aqueous solutions.
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X. Ding, P. A. Heiden
12
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continued for 3h at room temperature. The product was
collected onamagnet andwashedwith a3%v/v solutionof
acetic acid containing 0.1% w/v SDS. TEM images of the
protein-imprinted MIPNP show a diameter of �100nm
with an outer layer of �10nm. The authors reported that
the thickness of the imprinted layer could be controlled by
the DA self-polymerization time. Binding experiments
showed that the MIPNPs were highly selective to human
hemoglobin. The ease of producing theseMIPNPs using the
self-polymerizing DAmakes this an attractive approach for
imprinting suitable proteins.
Interestingly, Lee et al.[103] reported a simple method to
fabricate active enzyme (amylase) imprintedmagnetic NPs
with an average size of �100nm by a phase inversion
method of poly(ethylene-co-vinyl alcohol) (27–44 mol% of
ethylene) solutions in the presence of amylase templates.
Theamylase remainedactive for50cycles, asestablishedby
measuring glucose production from starch hydrolysis. By
this method, amylase-imprinted MIPNPs were prepared
that possessed high surface area, were easily separated
from the reaction mixture, and afforded rapid enzyme
reloading. MIPNPs that combine the superparamagnetism
of Fe3O4 with high selectivity and affinity to target
molecules are expected to be a valuable therapeutic and
biodiagnostic tool. Theymay also find value as amethod to
clear cytotoxic peptides from the bloodstream, as reported
by Hoshino et al.[112] Also, unlike other MIPNPs, an
applied external magnetic field might be used to guide
these MIPNPs to desired locations for action, and then aid
removal from the body. This alleviates the risk of NP
accumulation in the liver if theyarenoteffectively removed
from the body by the mononuclear phagocytic system.
3.2. Protein-Imprinted MIPNPs on Silica NP
Substrates
Most protein imprinted silica NPs are intended for use as
biosensorsor for solid-phaseextraction.Althoughmagnetic
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MIPNPs are more popular for protein separation, several
papers[104–106] reported the use of protein-imprinted silica
NPs for protein separation in solid phase extraction. Such
papers are representative of much of the literature on
designing protein-imprinted MIPNPs.
He et al.[104] imprinted lysozyme onto vinyl-functional-
ized silica NPs via radical polymerization using a low
concentration (0.4wt%) of functional monomers, which is
less than one-tenth of the concentration that is typically
used. The advantage of this low concentration is that
the possible aggregation of MIPNPs is avoided. The well-
dispersed MIPNP system allows a very thin protein-
imprinted film formed on the silica NPs surface, which
wasnotvisiblebyTEM.However, rebindingstudies showed
that the adsorption equilibriumwas achievedwithin 5min
with specific recognition towards the target protein.
This result indicates of importance of vinyl density on
the NP surface, when imprinting with vinyl monomers, on
the thickness of the imprinted layer and the binding site
density. This is corroborated by the study reported by Lu
et al.[45]. They changed the vinyl group spacing on the silica
NP surface, through control of the polymerization con-
ditions, and compared the thickness of the imprinted shell.
Therefore, both the shell thickness and density of binding
sites can be controlled in this way to achieve efficient and
accessible surface binding sites on MIPNPs.
3.3. Protein-Imprinted QD-Based MIPNPs
QD-based MIPNPs are a new type of chemo/biosensor
for protein detection. They combine the merits of the
optoelectronic properties of QDs with the recognition
specificity of MIPNPs. The principle of QD-based MIPNPs
was explained in the small molecule imprinting section. A
surface imprinting technique can also be used to fabricate
protein-imprinted QD-MIPNPs with excellent conforma-
tional integrity of the template protein and rapid rebinding
kinetics to target proteins. Different surface-modified
QDs have been developed to imprint different proteins.
For example, Tang et al.[107] synthesized bovine serum
albumin (BSA)-imprinted QD-MIP nanocomposites using
L-cysteine modified CdS QDs and a radical polymerization.
SEM images showed that the synthesized QD-MIPNPs
were aggregates with the imprinted layer coated on the
QD nanocrystals. After rebinding of the template BSA,
photoluminescence emission of QDs was quenched due
to fluorescence resonance energy transfer between QDs
and template. Zhang et al.[108] used a denatured bovine
serum albumin (dBSA) modified CdTe QDs and sol–gel
process to perform surface imprinting process. They used
APTES as a functional monomer and TEOS as a cross-linker
to form dBSA-imprinted QD-MIPNPs with particle sizes
of 30–50nm. Imprinted layers were formed on slightly
aggregated QDs.
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Lin et al.[109] reported a simple method to prepare
protein-imprinted QD-MIP nanocomposites by a phase
inversion technique. The host polymer solution (poly
(ethylene-co-vinyl alcohol)/DMSO solution) was mixed
together with a suitable amount of commercially available
QDs and template proteins, and the mixture was then
dispersed into non-solvent solution, such as deionized
water/isopropanol (2:3w/w), to form protein-imprinted
nanocomposites. Using this phase inversion technique,
they prepared creatinine-, albumin-, and lysozyme-
imprinted QD-MIP nanocomposites with particle size
ranging from 140 to 255nm. That study demonstrated
that the particle size of protein-imprinted QD-MIP nano-
composites and their morphologies were significantly
affected by reaction temperature when dispersing the
polymer/DMSO host solution in non-solvent. After remov-
ing the template proteins by dialysis, in 1wt% SDS solution
(15min) and deionized water several times, all three
protein-imprinted QD-MIP nanocomposites were used to
test real urine samples and showed good selective
recognition to the target proteins of creatinine, albumin,
and lysozyme, and possessed good accuracy.
Two different types of surface-modified QDs and one
convenient technique have been discussed above on the
fabrication of protein-imprinted QD-MIPNPs. Which type of
surface-modifiedQDsandmethods are suitable to fabricate a
specific protein-imprinted QD-MIPNP is mainly dependent
on theproperty of the template protein. In combinationwith
the techniques shown in small-molecularly imprinted QD-
MIPNPs, diverse techniques have been proposed to modify
QD surface functionalities, which make QD-MIPNPs a
promising platform for protein recognition and detection.
3.4. Protein-Imprinted MIPNPs by Precipitation
Polymerization
Precipitation polymerization is well-established polymeri-
zation method, and some recent advances in protein-
imprinted MIPNPs have been reported using this
method. The advances are achieved by reducing particle
size to enrich binding sites on/near the MIPNP surface
with easy accessibility for targets.[110,111] The examples
described in this section are selected to illustrate the
key factors for the protein imprinting process by precipita-
tion polymerization, including forming particles of a
suitable size and with good imprinting efficiency. To
accomplish this, mild reaction conditions in aqueous
solution are usually required, alongwith suitable function-
al monomers at a ratio that is empirically determined for
the specific template protein. These conditions and
procedures are usually required to avoid denaturization
of the protein, as well as to form effective interactions
between the functionalmonomers andappropriate regions
of the template proteins during the imprinting process.
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For example,Wang et al.[110] used precipitation polymer-
ization to prepare a MIPNP for specific adsorption of atrial
natriuretic peptide (ANP). The particle size was measured
at 215.8� 4.6 nm. The NH2–SLRRSS–CONH2 used as the
template peptide, which is a short peptide fromANP,MAA,
and NIPAAm as functional monomers, and MBAAm as
cross-linker. The optimal ratio of these reagents was
reported as MAA:NIPAAm:MBAAm at 1:3:10mol/mol.
The precipitation polymerization was carried out at room
temperature in aqueous media, with the assistance of a
small amount of surfactant (SDS), and initiated by
ammonium persulfate (APS) and N,N,N0, N0-tetramethyl-
ethylenediamine (TMED). The template peptide was
removed using 10wt% of acetic acid solution and washing
several times, leaving the recognition sites for both
template peptide and ANP. The binding kinetics test
revealed that MIPNPs reached protein-adsorption equilib-
rium within 30min, and showed a binding capacity of
106.7mmol g�1 for the template peptide and 36.0mmol g�1
for ANP, but showed little affinity to BSA or scrambledANP.
In 2010, Hoshino et al.[112] reported a type of MIPNP by
precipitation polymerization in mild conditions designed
to neutralize and clear cytotoxic target peptides (melittin)
from the bloodstream, functioning in the same way as
natural antibodies. The precipitation polymerization was
conducted inaqueous solutionwithvery lowconcentration
of surfactant at room temperature using an optimized
composition of functional monomers to yield melittin-
imprinted MIPNPs with a size distribution of 10–100nm.
In thiswork, they studied the effect of functionalmonomer
types and monomer ratio on the MIPNP size and yield.
When the functionalmonomerswere tert-butyl acrylamide
(TBAm) and acrylamide (AAm), used as a mole ratio of
40:5 the MIPNPs were 63nm and were obtained in 81%
yield. Similarly, when acrylic acid (AAc) was used instead
of AAm, still at the same 40:5, MIPNP was a 54nm and
obtained in 79% yield. When the researchers used TBAm,
AAm, and AAc at a mole ratio of 40:5:5, a 56nm MIPNP
was obtained in 88% yield.
Considering that template proteins possess complex
structures with hydrophobic, hydrophilic, and ionic
domains, a combination of the functional monomers
TBAm, AAm, and AAc (40:5:5 molar ratio) was chosen
that could provide hydrophobic interactions, hydrogen
bonding and electrostatic interactions with template
proteins, to yield effective recognition sites.[113] These
MIPNPs were termed ‘‘plastic antibodies’’ and expected
to perform in a manner similar to natural antibodies.
They were tested in the bloodstream of living mice with
effective adsorption of the toxin (melittin) and finally
accumulated in the liver to be expelled from the body.
This is the first report of MIPNPs being used as artificial
antibodies in the bloodstream. This interesting result
opens up new and promising applications for MIPNPs.
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4. Conclusion
The subject of MIPNPs is advancing rapidly. This short
review summarized recent progress in the area of MIPNPs,
with a focus on various surface imprinting techniques to
design MIPNPs using different NP substrates, such as silica
NPs, magnetic NPs, Au NPs, and QDs, and described some
of the advantages and applications of these MIPNPs.
Both small molecule and macromolecule imprinting were
described to show factors favoring binding sites formation,
accessibility, specific affinity to target molecules and fast
rebinding kinetics. The advantages of different synthetic
methods for the preparation of MIPNPs were illustrated
from literature examples, along with factors affecting
the surface imprinting process. This review also addressed
recent advances in designing MIPNPs to possess multiple
capabilities, such as combining magnetic responsiveness
withQDs or SPR properties into a singleMIPNP. Significant
advances have already been made that improve control
over surface imprinting, increase and allow MIPNPs to be
prepared that possess multi-functionality and can per-
form specific recognition, separation, and detection for
complex samples. It is expected that efforts to improve
specificity and impart multiple capability to MIPNPs
will continue, along with efforts to allow these MIPNPs
to be mass produced using cost-effective methods to
provide high throughput analyses to meet market
requirements.
Acknowledgements: The authors thank the Department ofChemistry at Michigan Technological University for financialsupport during the writing of this review article.
Received: April 14, 2013; Revised: June 16, 2013; Published online:DOI: 10.1002/mame.201300160
Keywords: biosensor; molecular imprinting; molecular recogni-tion; nanoparticles; surface imprinting
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