AC selected paper 201106~201205
1. "Label-Free Solution-Based Kinetic Study of Aptamer–Small Molecule Interactions by Kinetic Capillary
Electrophoresis with UV Detection Revealing How Kinetics Control Equilibrium." Anal. Chem. 2011, 83
(22), 8387-8390.
Here we demonstrate a label-free solution-based approach for studying the kinetics of biopolymer-small molecule
interactions. The approach utilizes kinetic capillary electrophoresis (KCE) separation and UV light absorption detection of the
unlabeled small molecule. In this proof-of-concept work, we applied KCE-UV to study kinetics of interaction between a small
molecule and a DNA aptamer. From the kinetic analysis of a series of aptamers, we found that dissociation rather than binding
controls the stability of the complex. Because of its label-free features and generic nature, KCE-UV promises to become a
practical tool for challenging kinetic studies of biopolymer-small molecule interactions.
2. "Continuous Signal Enhancement for Sensitive Aptamer Affinity Probe Electrophoresis Assay Using
Electrokinetic Concentration." Anal. Chem. 2011, 83 (18), 7086-7093.
We describe an electrokinetic concentration-enhanced aptamer affinity probe electrophoresis assay to achieve highly
sensitive and quantitative detection of protein targets in a microfluidic device. The key weaknesses of aptamer as a binding agent
(weak binding strength/fast target dissociation) were counteracted by continuous injection of fresh sample while band-broadening
phenomena were minimized due to self-focusing effects. With 30 min of continuous signal enhancement, we can detect 4.4 pM
human immunoglobulin E (IgE) and 9 pM human immunodeficiency virus 1 reverse transcriptase (HIV-1 RT), which are among
the lowest limits of detection (LOD) reported. IgE was detected in serum sample with a LOD of 39 pM due to nonspecific
interactions between aptamers and serum proteins. The method presented in this paper also has broad applicability to improve
sensitivities of various other mobility shift assays.
3. "Pressure-Based Approach for the Analysis of Protein Adsorption in Capillary Electrophoresis." Anal.
Chem. 2011, 84 (1), 453-458.
Protein adsorption to inner capillary walls creates a major obstacle in all applications of capillary electrophoresis involving
protein samples. The problem is especially severe in kinetic capillary electrophoresis (KCE) techniques, which are used to study
protein ligand interactions at physiological conditions and, thus, cannot utilize extreme pH. A variety of coatings exist to reduce
protein adsorption in CE, each expressing a unique surface chemistry that interacts with individual proteins differently. Here we
introduce a simple pressure-based method for the qualitative assessment of protein adsorption that can facilitate the direct
antiadhesive ranking of several coatings toward a protein of interest. In this approach, a short plug of the protein is injected into a
capillary and propagated through with a pressure low enough to ensure adequate Taylor dispersion. The experiment is performed
with a nonmodified commercial instrument in a pseudo-two-detector approach. The two detectors are mimicked by using two
different distances from the capillary inlet to a single detector. If the peak area and shape do not change with changing distance,
the protein does not adsorb appreciably, while a decreasing peak area with increasing distance infers inner surface adsorption. The
magnitude change of the peak area between the two distances along with the overall peak shape is used to gauge the extent of
protein adsorption. By using this method, we ranked antiadhesive properties of different wall chemistries for a series of proteins.
The described method will be useful for optimizing protein analysis by CE and, in particular, for KCE experiments that
investigate how proteins interact with their respective ligands.
4. "Peak-Shape Correction to Symmetry for Pressure-Driven Sample Injection in Capillary Electrophoresis."
Anal. Chem. 2011, 84 (1), 149-154.
Pressure-driven sample injection in capillary electrophoresis results in asymmetric peaks due to difference in shapes between
the front and the back boundaries of the sample plug. Uneven velocity profile of fluid flow across the capillary gives the front
boundary a parabolic shape. The back side, on the other hand, has a flat interface with the electrophoresis run buffer. Here, we
propose a simple means of correcting this asymmetry by pressure-driven "propagation" of the injected plug, with the parabolic
sample-buffer interface established at the back. We prove experimentally that such a propagation procedure corrects peak
asymmetry to the level comparable to injection through electroosmosis. Importantly, the propagation-based correction procedure
also solves a problem of transferring the sample into the efficiently cooled zone of the capillary for capillary electrophoresis (CE)
instruments with active cooling. The suggested peak correction procedure will find applications in all CE methods that rely on
peak shape analysis, e.g., nonequilibrium capillary electrophoresis of equilibrium mixtures.
5. "Distance-Dependent Metal-Enhanced Quantum Dots Fluorescence Analysis in Solution by Capillary
Electrophoresis and Its Application to DNA Detection." Anal. Chem. 2011, 83 (11), 4103-4109.
Here the distance dependence of metal-enhanced quantum dots (QDs) fluorescence in solution is studied systematically by
capillary electrophoresis (CE). Complementary DNA oligonucleotides-modified CdSe/ZnS QDs and gold nanoparticles (Au NPs)
were connected together in solution by the hybridization of complementary oligonucleotides, and a model system (QD-Au) for the
study of metal-enhanced QDs fluorescence was constructed, in which the distance between the QDs and Au NPs was controlled
by adjusting the base number of the oligonucleotide. In our CE experiments, the metal-enhanced fluorescence of the QDs solution
was only observed when the distance between the QDs and Au NPs ranged from 6.8 to 18.7 nm, and the maximum enhancement
by a factor of 2.3 was achieved at 11.9 nm. Furthermore, a minimum of 19.6 pg of target DNA was identified in CE based on its
specific competition with the QD-DNA in the QD-Au system. This work provides an important reference for future study of
metal-enhanced QDs fluorescence in solution and exhibits potential capability in nucleic acid hybridization analysis and
high-sensitivity DNA detection.
6. "Simultaneous Electrokinetic and Hydrodynamic Injection for High Sensitivity Bacteria Analysis in
Capillary Electrophoresis." Anal. Chem. 2011, 83 (12), 4949-4954.
A repeatable preconcentration electrophoretic methodology for the analysis of bacteria was developed. This method is based
on an isotachophoretic mode coupled with a simultaneous hydrodynamic-electrokinetic injection in conditions of field-amplified
sample injection. This electrophoretic method allows the quantification of Enterobacter cloacae (studied as a model of Gram
negative bacteria) with a limit of detection of 2 x 10(4) cells/mL. With the optimized conditions, a preconcentration factor of
about 500-fold was obtained as compared to a standard hydrodynamic injection. The RSD (n = 5) on the migration time and on
the peak area were 3% and 5%, respectively. This capillary electrophoretic methodology has been applied for the quantification of
microbes in natural water (liver and natural spring waters). Filtration of the sample prior to injection was required to remove ions
present in the water and to keep the field-amplified sample injection condition at the injection. Filtrated bacteria were then
recovered in terminating electrolyte diluted 10 times with water. Good agreements were obtained between cellular ATP
measurements and the proposed CE methodology for the quantification of bacteria in waters..
7. "High-Resolution Separation of Graphene Oxide by Capillary Electrophoresis." Anal. Chem. 2011, 83
(23), 9100-9106.
Separation and purification of graphene oxide (GO) prepared from chemical oxidation of flake graphite and ultrasonication
by capillary electrophoresis (CE) was demonstrated. CE showed the ability to provide high-resolution separations of GO
fractionations with baseline separation. The GO fractionations after CE were collected for Raman spectroscopy, atomic force
microscopy, and transmission electron microscopy characterizations. GO nanoparticles (unexfoliated GO) or stacked GO sheets
migrated toward the anode, while the thin-layer GO sheets migrated toward the cathode. Therefore, CE has to be performed twice
with a reversed electric field to achieve a full separation of GO. This separation method was suggested to be based on the surface
charge of the GO sheets, and a separation model was proposed. This study might be valuable for fabrication of GO or graphene
micro- or nanodevices with controlled thickness.
8. "Study of Antibacterial Activity by Capillary Electrophoresis Using Multiple UV Detection Points." Anal.
Chem. 2012, 84 (7), 3302-3310.
A new methodology for an antibacterial assay based on capillary electrophoresis with multiple UV detection points has been
proposed. The possible antibacterial activity of cationic molecules on bacteria (Gram-positive and Gram-negative) is studied by
detecting the bacteria before, during, and after their meeting with the cationic antibacterial compound. For that, a UV area
imaging detector having two loops and three detection windows was used with a 95 cm x 100 mu m i.d. capillary. In the
antibacterial assay, the bacteria (negatively charged) and the cationic molecules were injected separately from each end of the
capillary. The bacteria were mobilized by anionic ITP mode while cationic molecules migrate in the opposite direction under
conditions close to CZE. The cationic molecules were injected into the capillary as a broad band (injected volume about 16% of
the volume of the capillary) to prevent dilution of the sample during the electrophoretic process. Bacteriolytic activity, as well as
strong interactions between the small antibacterial molecules and the bacteria, can be investigated within a few minutes. The assay
was used to study the antibacterial activity of dendrigraft poly-L-lysines on Micrococcus luteus and Erwinia carotovora. Because
dendrigraft poly-L-lysines are nonimmunogenic and have low toxicity, this new class of dendritic biomacromolecules is very
promising for antibacterial applications.
9. "Facile Preparation of Graphene-Copper Nanoparticle Composite by in Situ Chemical Reduction for
Electrochemical Sensing of Carbohydrates." Anal. Chem. 2011, 84 (1), 171-178.
A novel graphene-copper nanoparticle composite was prepared by the in situ chemical reduction of a mixture containing
graphene oxide and copper(II) ions using potassium borohydride as a reductant. It was mixed with paraffin oil and packed into
one end of a fused capillary to fabricate microdisc electrodes for sensing carbohydrates. The morphology and structure of the
graphene-copper nanoparticle composite were investigated by scanning electron microscopy, X-ray diffraction, and Fourier
transform-infrared spectroscopy. The results indicated that copper nano-particles with an average diameter of 20.8 nm were
successfully deposited on graphene nanosheets to form a well interconnected hybrid network The analytical performance of these
unique graphene-copper nanoparticle composite paste electrodes was demonstrated by sensing five carbohydrates in combination
with cyclic voltammetry and capillary electrophoresis (CE). The advantages of the composite detectors include higher sensitivity,
satisfactory stability, surface renewability, bulk modification, and low expense of fabrication. They should find applications in
microchip CE, flowing-injection analysis, and other microfluidic analysis systems.
10. "Recyclable and High-Sensitivity Electrochemical Biosensing Platform Composed of Carbon-Doped
TiO2 Nanotube Arrays." Anal. Chem. 2011, 83 (21), 8138-8144.
Electrode fouling and passivation are the main reasons for attenuated signals as well as reduced sensitivity and selectivity
over time in electrochemical analysis. We report here a refreshable electrode composed of carbon-doped TiO2 nanotube arrays
(C-doped TiO2-NTAs), which not only has excellent electrochemical activity for simultaneous determination of
5-hydroxytryptamine and ascorbic acid but also can be easily photocatalytically refreshed to maintain the high selectivity and
sensitivity. The C-doped TiO2-NTAs are fabricated by rapid annealing of as-anodized TiO2-NTAs in argon. The residual ethylene
glycol absorbed on the nanotube wall acts as the carbon source and no foreign carbon precursor is thus needed. The morphology,
structure, and composition the C-doped TiO2-NTAs are determined, and the corresponding doping mechanism is investigated by
thermal analysis and in situ mass spectroscopy. Because of the high photocatalytic activity of the C-doped TiO2-NTAs electrode,
the electrode surface can be readily regenerated by ultraviolet or visible light irradiation. This photoassisted regenerating
technique does not damage the electrode microstructure while rendering high reproducibility and stability.
11. "In-Channel Electrochemical Detection in the Middle of Microchannel under High Electric Field." Anal.
Chem. 2011, 84 (2), 901-907.
We propose a new method for performing in-channel electrochemical detection under a high electric field using a
polyelectrolytic gel salt bridge (PGSB) integrated in the middle of the electrophoretic separation channel. The finely tuned
placement of a gold working electrode and the PGSB on an equipotential surface in the microchannel provided highly sensitive
electrochemical detection without any deterioration in the separation efficiency or interference of the applied electric field. To
assess the working principle, the open circuit potentials between gold working electrodes and the reference electrode at varying
distances were measured in the microchannel under electrophoretic fields using an electrically isolated potentiostat. In addition,
"in-channel" cyclic voltammetry confirmed the feasibility of electrochemical detection under various strengths of electric fields
(similar to 400 V/cm). Effective separation on a microchip equipped with a PGSB under high electric fields was demonstrated for
the electrochemical detection of biological compounds such as dopamine and catechol. The proposed "in-channel"
electrochemical detection under a high electric field enables wider electrochemical detection applications in microchip
electrophoresis.
12. "Electrochemical Sensing of Aptamer-Facilitated Virus Immunoshielding." Anal. Chem. 2011, 84 (3),
1677-1686.
Oncolytic viruses (OVs) are promising therapeutics that selectively replicate in and kill tumor cells. However, repetitive
administration of OVs provokes the generation of neutralizing antibodies (nAbs) that can diminish their anticancer effects. In this
work, we selected DNA aptamers against an oncolytic virus, vesicular stomatitis virus (VSV), to protect it from nAbs. A
label-free electrochemical aptasensor was used to evaluate the degree of protection (DoP). The aptasensor was fabricated by
self-assembling a hybrid of a thiolated ssDNA primer and a VSV-specific aptamer. Electrochemical impedance spectroscopy was
employed to quantitate VSV in the range of 800-2200 PFU and a detection limit of 600 PFU. The aptasensor was also utilized for
evaluating binding affinities between VSV and aptamer pools/clones. An electrochemical displacement assay was performed in
the presence of nAbs and DoP values were calculated for several VSV-aptamer pools/clones. A parallel flow cytometric analysis
confirmed the electrochemical results. Finally, four VSV-specific aptamer clones, ZMYK-20, ZMYK-22, ZMYK-23, and
ZMYK-28, showed the highest protective properties with dissociation constants of 17, 8, 20, and 13 nM, respectively. Another
four sequences, ZMYK-1, -21, -25, and -29, exhibited high affinities to VSV without protecting it from nAbs and can be further
utilized in sandwich assays. Thus, ZMYK-22, -23, and -28 have the potential to allow efficient delivery of VSV through the
bloodstream without compromising the patient's immune system.
13. "DNA Nanostructure-Decorated Surfaces for Enhanced Aptamer-Target Binding and Electrochemical
Cocaine Sensors." Anal. Chem. 2011, 83 (19), 7418-7423.
The sensitivity of aptamer-based electrochemical sensors is often limited by restricted target accessibility and
surface-induced perturbation of the aptamer structure, which arise from imperfect packing of probes on the heterogeneous and
locally crowded surface. In this study, we have developed an ultrasensitive and highly selective electrochemical aptamer-based
cocaine sensor (EACS), based on a DNA nanotechnology-based sensing platform. We have found that the electrode surface
decorated with an aptamer probe-pendant tetrahedral DNA nanostructure greatly facilitates cocaine-induced fusion of the split
anticocaine aptamer. This novel design leads to a sensitive cocaine sensor with a remarkably low detection limit of 33 nM. It is
also important that the tetrahedra-decorated surface is protein-resistant, which not only suits the enzyme-based signal
amplification scheme employed in this work, but ensures high selectivity of this sensor when deployed in sera or other adulterated
samples.
14. "Rational Design and One-Step Formation of Multifunctional Gel Transducer for Simple Fabrication of
Integrated Electrochemical Biosensors." Anal. Chem. 2011, 83 (14), 5715-5720.
This study demonstrates a new strategy to simplify the biosensor fabrication and thus minimize the biosensor-to-biosensor
deviation through rational design and one-step formation of a multifunctional gel electronic transducer integrating all elements
necessitated for efficiently transducing the biorecognition events to signal readout, by using glucose dehydrogenase (GDH) based
electrochemical biosensor as an example. To meet the requirements for preparing integrated biosensors and retaining electronic
and ionic conductivities for electronically transducing process, ionic liquids (ILs) with enzyme cofactor (i.e., oxidized form of
nicotinamide adenine dinucleotide) as the anion were synthesized and used to form a bucky gel with single-walled carbon
nanotubes, in which methylene green electrocatalyst was stably encapsulated for the oxidation of nicotinamide adenine
dinucleotide. With such kind of rationally designed and one-step-formed multifunctional gel as the electronic transducer, the
GDH-based electrochemical biosensors were simply fabricated by polishing the electrodes onto the gel followed by enzyme
immobilization. This capability greatly simplifies the biosensor fabrication, prolongs the stability of the biosensors, and, more
remarkably, minimizes the biosensor-to-biosensor deviation. The relative standard deviations obtained both with one electrode for
the repeated measurements of glucose and with the different electrodes prepared with the same method for the concurrent
measurements of glucose with the same concentration were 3.30% (n = 7) and 4.70% (n = 6), respectively. These excellent
properties of the multifunctional gel-based biosensors substantially enable them to well-satisfy the pressing need of rapid
measurements, for example, environmental monitoring, food analysis, and clinical diagnoses.
15. "An Electrochemically Reduced Graphene Oxide-Based Electrochemical Immunosensing Platform for
Ultrasensitive Antigen Detection." Anal. Chem. 2012, 84 (4), 1871-1878.
We present an electrochemically reduced graphene oxide (ERGO)-based electrochemical immunosensing platform for the
ultrasensitive detection of an antigen by the sandwich enzyme-linked immunosorbent assay (ELISA) protocol. Graphene oxide
(GO) sheets were initially deposited on the amine-terminated benzenediazonium-modified indiun tin oxide (ITO) surfaces through
both electrostatic and pi-pi interactions between the modified surfaces and GO. This deposition was followed by the
electrochemical reduction of graphene oxide (GO) for preparing ERGO-modified ITO surfaces. These surfaces were then coated
with an N-acryloxysuccinimide-activated amphiphilic polymer, poly(BMA-r-PEGMA-r-NAS), through pi-pi stacking interactions
between the benzene ring tethered to the polymer and ERGO. After covalent immobilization of a primary antibody on the
polymer-modified surfaces, sandwich ELISA was carried out for the detection of an antigen by use of a horseradish peroxidase
(HRP)-labeled secondary antibody. Under the optimized experimental conditions, the developed electrochemical immunosensor
exhibited a linear response over a wide range of antigen concentrations with a very low limit of detection (ca. 100 fg/mL, which
corresponds to ca. 700 aM). The high sensitivity of the electrochemical immunosensor may be attributed not only to the enhanced
electrocatalytic activity owing to ERGO but also to the minimized background current owing to the reduced nonspecific binding
of proteins.
16. "Electrochemical Aptamer-Based Sandwich Assays for the Detection of Explosives." Anal. Chem. 2012,
84 (10), 4245-4247.
Electrochemical impedance spectroscopy (EIS) is used to detect 2,4,6-trinitrotoluene (TNT) in a novel sandwiched structure
which relies on the specific interactions between (i) primary amine with TNT and (ii) TNT and anti-TNT aptamer. With pure
targets, the assay has a sensitivity of 10–14 M, a dynamic range of 10–14–10–3 M, and employs a small sample volume (25 μL).
The method’s sensitivity is comparable to state of the art optical methods with the added advantages of electrochemical detection,
which can be easily miniaturized and implemented into a hand-held device.
17. "Probing DNA’s Interstrand Orientation with Gold Nanoparticles." Anal. Chem. 2011, 83 (13),
5067-5072.
The interstrand orientation of a DNA duplex plays a pivotal role in its biological and chemical functions. Therefore,
developing an efficient way to determine (control and monitor) the parallel or antiparallel conformation of a DNA duplex is of
great significance, which, however, remains a big challenge under some circumstances. In this work, we demonstrate that gold
nanoparticles tagged on DNA are especially useful in trapping and detecting a special interstrand orientation of a DNA double
helix, based on inherent electrostatic and steric repulsions between nanoparticles which will affect their self-assembly into a large
structure. More importantly, some of the conformations revealed by the gold nanoparticle assay may even not be
thermodynamically preferred and thus will be hard to detect using currently available methods. This simple, straightforward, and
efficient methodology capable of dictating and probing a special DNA duplex structure provides a useful tool for conformational
analyses and functional explorations of biomolecules as well as biophysical and nanobiomedical research.
18. "Fluorescent Ferritin Nanoparticles and Application to the Aptamer Sensor." Anal. Chem. 2011, 83 (15),
5834-5843.
We synthesized fluorescent ferritin nanoparticles (FFNPs) through bacterial expression of the hybrid gene consisting of
human ferritin heavy chain (hFTN-H), spacer (glycine-rich peptide), and enhanced green (or red) fluorescent protein [eGFP (or
DsRed)] genes. The self-assembly activity of hFTN-H that leads to the formation of nanoparticles (12 nm in diameter), the
conformational flexibility of the C-terminus of hFTN-H, and the glycine-rich spacer enabled eGFPs (or DsReds) to be well
displayed on the surface of each ferritin nanoparticle, resulting in the construction of green (or red) FFNPs [gFFNPs (or rFFNPs)].
As compared to eGFP (or DsRed) alone, it is notable that the developed FFNPs showed significantly amplified fluorescence
intensity and also enhanced stability. DNA aptamers were chemically conjugated to gFFNP via each eGFP's cysteine residue that
was newly introduced through site-directed mutagenesis (Ser175Cys). The DNA-aptamer-conjugated gFFNPs were used as a
fluorescent reporter probe in the aptamer-based "sandwich" assay of a cancer marker [i.e., platelet-derived growth factor B-chain
homodimer (PDGF-BB)] in phosphate-buffered saline buffer or diluted human serum. This is a simple two-step assay without any
additional steps for signal amplification, showing that compared to the same aptamer-based assays using eGFP alone or Cy3, the
detection signals, affinity of the reporter probe to the cancer marker, and assay sensitivity were significantly enhanced; i.e., the
limit of detection was lowered to the 100 fM level. Although the PDGF-BB assay is reported here as a proof-of-concept, the
developed FFNPs can be applied in general to any aptamer-based sandwich assays.
19. "Fluorescence Enhancement of Silver Nanoparticle Hybrid Probes and Ultrasensitive Detection of IgE."
Anal. Chem. 2011, 83 (23), 8945-8952.
An ultrasensitive protein assay method was developed based on silver nanoparticle (AgNP) hybrid probes and
metal-enhanced fluorescence. Two aptamer based silver nanoparticles, Aptamer/Oligomer-A/Cy3-modified AgNPs (Tag-A) and
Aptamer/Oligomer-B/Cy3-modified AgNPs (Tag-B) were hybridized to form a silver nanoparticle aggregate that produced a red
shift and broadening of the Localized Surface Plasmon Resonance (LSPR) peak. The enhanced fluorescence resulted from the
increased content of Cy3 molecules and their emission resonance coupled to the broadened localized surface plasmon (LSP) of
AgNP aggregate. The separation distance between Cy3 and AgNPs was 8 nm which was the most optimal for metal enhanced
fluorescence and the separation distance between adjacent AgNPs was about 16 nm and this was controlled by the lengths of
oligomer-A and oligomer-B. The protein array was prepared by covalently immobilizing capture antibodies on aldehyde-coated
slide. After addition of protein IgE sample, two kinds of aptamer-modified AgNPs (Tag-A and Tag-B) were employed to
specifically recognize IgE and form the AgNP aggregate on the arrays based on their hybridization. The detection property of the
aptamer-modified AgNP aggregate was compared to two other modified aptamer-based probes, aptamer-modified Cy3 and Tag-A.
The modified AgNP hybrid probe (Tag-A and Tag-B) showed remarkable superiority in both sensitivity and detection limit due to
the formed AgNP aggregate. The new hybrid probe also produced a wider linear range from 0.49 to 1000 ng/mL. with the
detection limit reduced to 40 pg/mL (211 fM). The presented method showed that the newly designed strategy of combining
aptamer-based nanomaterials to form aggregates results in a highly sensitive optical detection method based on localized surface
plasmon.
20. "Nanoparticle PEBBLE Sensors for Quantitative Nanomolar Imaging of Intracellular Free Calcium
Ions." Anal. Chem. 2011, 84 (2), 978-986.
Ca2+ is a universal second messenger and plays a major role in intracellular signaling, metabolism, and a wide range of
cellular processes. To date, one of the most successful approaches for intracellular Ca2+ measurement involves the introduction of
optically sensitive Ca2+ indicators into living cells, combined with digital imaging microscopy. However, the use of free Ca2+
indicators for intracellular sensing and imaging has several limitations, such as nonratiometric measurement for the most-sensitive
indicators, cytotoxicity of the indicators, interference from nonspecific binding caused by cellular biomacromolecules, challenging
calibration, and unwanted sequestration of the indicator molecules. These problems are minimized when the Ca2+ indicators are
encapsulated inside porous and inert polyacrylamide nanoparticles. We present PEBBLE nanosensors encapsulated with
rhodamine-based Ca2+ fluorescence indicators. The rhod-2-containing PEBBLEs presented here show a stable sensing range at
near-neutral pH (pH 6-9). Because of the protection of the PEBBLE matrix, the interference of protein-nonspecific binding to the
indicator is minimal. The rhod-2 PEBBLEs give a nanomolar dynamic sensing range for both in-solution (K-d = 478 nM) and
intracellular (K-d = 293 nM) measurements. These nanosensors are useful quantitative tools for the measurement and imaging of
the cytosolic nanomolar free Ca2+ levels.
21. "Bacterial Inactivation Using Silver-Coated Magnetic Nanoparticles as Functional Antimicrobial
Agents." Anal. Chem. 2011, 83 (22), 8688-8695.
The ability for silver nanoparticles to function as an antibacterial agent while being separable from the target fluids is
important for bacterial inactivation in biological fluids. This report describes the analysis of the antimicrobial activities of
silver-coated magnetic nanoparticles synthesized by wet chemical methods. The bacterial inactivation of several types of bacteria
was analyzed, including Gram-positive bacteria (Staphylococcus aureus and Bacillus cereus) and Gram-negative bacteria
(Pseudomonas aeruginosa, Enterobacter cloacae, and Escherichia coli). The results have demonstrated the viability of the
silver-coated magnetic nanoparticles for achieving effective bacterial inactivation efficiency comparable to and better than that of
silver nanoparticles conventionally used. The bacteria inactivation efficiency of our silver-coated MnZn ferrite (MZF@Ag)
nanoparticles was also determined for blood platelets samples, demonstrating the potential of utilization in inactivating bacterial
growth in platelets prior to transfusion to ensure blood product safety, which also has important implications for enabling the
capability of effective separation, delivery, and targeting of the antibacterial agents.
22. "Aptamer Capturing of Enzymes on Magnetic Beads to Enhance Assay Specificity and Sensitivity."
Anal. Chem. 2011, 83 (24), 9234-9236.
Activity and specificity of enzyme molecules are important to enzymatic reactions and enzyme assays. We describe an
aptamer capturing approach that improves the specificity and the sensitivity of enzyme detection. An aptamer recognizing the
target enzyme molecule is conjugated on a magnetic bead, increasing the local concentration, and serves as an affinity probe to
capture and separate minute amounts of the enzyme. The captured enzymes catalyze the subsequent conversion of fluorogenic
substrate to fluorescent products, enabling a sensitive measure of the active enzyme. The feasibility of this technique is
demonstrated through assays for human alpha thrombin and human neutrophil elastase (HNE), two important enzymes. Thrombin
(2 fM) and 100 fM HNE can be detected. The incorporation of two binding events, substrate recognition and aptamer binding,
greatly improves assay specificity. With its simplicity, this approach is applicable to biosensing and detection of disease
biomarkers.
23. "Aptamer-Mediated Nanoparticle-Based Protein Labeling Platform for Intracellular Imaging and
Tracking Endocytosis Dynamics." Anal. Chem. 2012, 84 (7), 3099-3110.
Although nanoparticles have been widely used as optical contrasts for cell imaging, the complicated prefunctionalized steps
and low labeling efficiency of nanoprobes greatly inhibit their applications in cellular protein imaging. In this study, we developed
a novel and general strategy that employs an aptamer not only as a recognizer for protein recognition but also as a linker for
nanoreporter targeting to specifically label membrane proteins of interest and track their endocytic pathway. With this strategy,
three kinds of nanoparticles, including gold nanoparticles, silver nanoparticles, and quantum dots (QDs), have been successfully
targeted to the membrane proteins of interest, such as nucleolin or prion protein (PrPC). The following investigations on the
subcellular distribution with fluorescent immunocolocalization assay indicated that PrPC-aptamer-QD complexes most likely
internalized into cytoplasm through a classical clathrin-dependent/receptor-mediated pathway. Further single-particle tracking and
trajectory analysis demonstrated that PrPC-aptamer-QD complexes exhibited a complex dynamic process, which involved three
types of movements, including membrane diffusion, vesicle transportation, and confined diffusion, and all types of these
movements were associated with distinct phases of PrPC endocytosis. Compared with traditional multilayer methods, our
proposed aptamer-mediated strategy is simple in procedure, avoiding any complicated probe premodification and purification. In
particular, the new double-color labeling strategy is unique and significant due to its superior advantages of targeting two signal
reporters simultaneously in a single protein using only one aptamer. What is more important, we have constructed a general and
versatile aptamer-mediated protein labeling nanoplatform that has shown great promise for future biomedical labeling and
intracellular protein dynamic analysis.
24. "Robust and Highly Sensitive Fluorescence Approach for Point-of-Care Virus Detection Based on
Immunomagnetic Separation." Anal. Chem. 2012, 84 (5), 2358-2365.
In this work, robust approach for a highly sensitive point-of-care virus detection was established based on immunomagnetic
nanobeads and fluorescent quantum dots (QDs). Taking advantage of immunomagnetic nanobeads functionalized with the
monoclonal antibody (mAb) to the surface protein hemagglutinin (HA) of avian influenza virus (AIV) H9N2 subtype, H9N2
viruses were efficiently captured through antibody affinity binding, without pretreatment of samples. The capture kinetics could
be fitted well with a first-order bimolecular reaction with a high capturing rate constant k(f) of 4.25 x 10(9) (mol/L)(-1) s(-1),
which suggested that the viruses could be quickly captured by the well-dispersed and comparable-size immunomagnetic
nanobeads. In order to improve the sensitivity, high-luminance QDs conjugated with streptavidin (QDs-SA) were introduced to
this assay through the high affinity biotin-streptavidin system by using the biotinylated mAb in an immuno sandwich mode. We
ensured the selective binding of QDs-SA to the available biotin-sites on biotinylated mAb and optimized the conditions to reduce
the nonspecific adsorption of QDs-SA to get a limit of detection low up to 60 copies of viruses in 200 mu L. This approach is
robust for application at the point-of-care due to its very good specificity, precision, and reproducibility with an intra-assay
variability of 1.35% and an interassay variability of 3.0%, as well as its high selectivity also demonstrated by analysis of synthetic
biological samples with mashed tissues and feces. Moreover, this method has been validated through a double-blind trial with 30
throat swab samples with a coincidence of 96.7% with the expected results.
25. "Light-Controlled Bioelectrochemical Sensor Based on CdSe/ZnS Quantum Dots." Anal. Chem. 2011,
83 (20), 7778-7785.
This study reports on the oxygen sensitivity of quantum dot electrodes modified with CdSe/ZnS nanocrystals. The
photocurrent behavior is analyzed for dependence on pH and applied potential by potentiostatic and potentiodynamic
measurements. On the basis of the influence of the oxygen content in solution on the photocurrent generation, the enzymatic
activity of glucose oxidase is evaluated in solution. In order to construct a photobioelectrochemical sensor which can be read out
by illuminating the respective electrode area, two different immobilization methods for the fixation of the biocatalyst have been
investigated. Both covalent cross-linking and layer-by-layer deposition of GOD by means of the polyelectrolyte polyallylamine
hydrochloride show that a sensor construction is possible. The sensing properties of this type of electrode are drastically
influenced by the amount and density of the enzyme on top of the quantum dot layer, which can be advantageously adjusted by
the layer-by-layer technique. By depositing four bilayers [GOD/PAH](4) on the CdSe/ZnS electrode, a fast-responding sensor for
the concentration range of 0.1-5 mM glucose can be prepared. This study opens the door to multianalyte detection with a
nonstructured sensing electrode, localized enzymes, and spatial read-out by light.