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1.
Lillian Roth Jutta Zagon Anke Ehlers Lothar W. Kroh Hermann Broll 《Analytical and bioanalytical chemistry》2009,394(2):529-537
A new approach for the detection of DNA target molecules is described, using capture probes and subsequent signal enhancement
by a uniform polymerase chain reaction (PCR). Peptide nucleic acid probes were immobilized in real-time PCR-compatible microtiter
plates. After hybridization of biotinylated DNA targets, detection was performed by real-time immuno-PCR, a method formerly
used for protein detection. We demonstrate the feasibility of this strategy for the qualitative detection of DNA oligonucleotides
with a detection limit (LOD) of 6 attomol. Furthermore, the method was applied to PCR-amplified samples from genetically modified
maize DNA (Mon810). A 483-bp DNA fragment was detected in mixture with 99.9% of noncomplementary DNA with a sensitivity down
to the level of attomole.
Figure 相似文献
2.
Addition of gold nanoparticles to real-time PCR: effect on PCR profile and SYBR Green I fluorescence
Haber AL Griffiths KR Jamting AK Emslie KR 《Analytical and bioanalytical chemistry》2008,392(5):887-896
Real-time quantitative polymerase chain reaction (qPCR) is the industry standard technique for the quantitative analysis of
nucleic acids due to its unmatched sensitivity and specificity. Optimisation and improvements of this fundamental technique
over the past decade have largely consisted of attempts to allow faster and more accurate ramping between critical temperatures
by improving assay reagents and the thermal geometry of the PCR chamber. Small gold nanoparticles (Au-NPs) have been reported
to improve PCR yield under fast cycling conditions. In this study, we investigated the effect of Au-NPs on optimised real-time
qPCR assays by amplifying DNA sequences from genetically modified canola in the presence and absence of 0.9 nM Au-NPs of diameter
12 ± 2nm. Contrary to expectations, we found that Au-NPs altered the PCR amplification profile when using a SYBR Green I detection
system due to fluorescence quenching; furthermore, high-resolution melt (HRM) analysis demonstrated that Au-NPs destabilised
the double-stranded PCR product. The results indicate that effects on the assay detection system must be carefully evaluated
before Au-NPs are included in any qPCR assay.
Figure Raw amplification profiles in the presence and absence of gold nanoparticles 相似文献
3.
The use of polymers in microchip fabrication affords new opportunities for the development of powerful, miniaturized separation
techniques. One method in particular, the use of phase-changing sacrificial layers, allows for simplified designs and many
additional features to the now standard fabrication of microchips. With the possibility of adding a third dimension to the
design of separation devices, various means of enhancing analysis now become possible. The application of phase-changing sacrificial
layers in microchip analysis systems is discussed, both in terms of current uses and future possibilities.
Figure Phase-changing sacrificial materials enable multilayer microfluidic device layouts 相似文献
4.
Single molecule detection in nanofluidic digital array enables accurate measurement of DNA copy number 总被引:3,自引:0,他引:3
Somanath Bhat Jan Herrmann Paul Armishaw Philippe Corbisier Kerry R. Emslie 《Analytical and bioanalytical chemistry》2009,394(2):457-467
Digital polymerase chain reaction (PCR) is a promising technique for estimating target DNA copy number. PCR solution is distributed
throughout numerous partitions, and following amplification, target DNA copy number is estimated based on the proportion of
partitions containing amplified DNA. Here, we identify approaches for obtaining reliable digital PCR data. Single molecule
amplification efficiency was significantly improved following fragmentation of total DNA and bias in copy number estimates
reduced by analysis of short intact target DNA fragments. Random and independent distribution of target DNA molecules throughout
partitions, which is critical to accurate digital PCR measurement, was demonstrated by spatial distribution analysis. The
estimated relative uncertainty for target DNA concentration was under 6% when analyzing five digital panels comprising 765
partitions each, provided the panels contained an average of 212 to 3,365 template molecules. Partition volume was a major
component of this uncertainty estimate. These findings can be applied to other digital PCR studies to improve confidence in
such measurements.
Figure Digital PCR amplification plot (left) and panel read out (right) of HindIII-digested pIRMM69. pIRMM69 contains one HindIII restriction enzyme site outside the hmg and transgene fragments used as targets in PCR. Red boxes with white shade denote positive hits containing one or more target
DNA molecules, and white boxes with grey shade refer to no target being amplified. 相似文献
5.
A novel method for rapid separation and determination of ascorbic acid and uric acid has been developed with a polycation-modified
poly(dimethylsiloxane) (PDMS) microchip under a negative-separation electric field. Just by flushing the microchip with aqueous
solutions of the polycations, poly(allylamine) hydrochloride, poly(diallyldimethylammonium chloride) or chitosan could be
stably coated on the PDMS microchannel surface, which resulted in a reversed electroosmotic flow and thus the rapid and efficient
separation of the two substrates. Factors influencing the separation, including polycation category, buffer solution, detection
potential and separation voltage, were investigated and optimized. The cheapness, rapid analysis speed and the successful
analysis of human urine make this microsystem attractive for application in clinics.
Figure The electropherograms of 100 μ/mL AA and UA in (1) PAH, (2) PDDA, (3) Chitosan modified PDMS microchannels and native PDMS
microchip (4). 相似文献
6.
Christoph Logé Alexander Bornschlegl Ulrich Boesl 《Analytical and bioanalytical chemistry》2009,395(6):1631-1639
Circular dichroism in ion yield has promising new potentials for chiral analysis. Our progress of its development is described
here. Circular dichroism in ion yield is achieved by resonance-enhanced multiphoton ionization. The feasibility of circular
dichroism spectroscopy and quantitative determination of circular dichroism by this method is demonstrated. Several excitation
schemes have been applied using different types of lasers, which vary in wavelength and repetition rate. Progress to improve
the statistical error and thus the lower limit of measurable circular dichroism is described. This is achieved by adding achiral
compounds or racemic mixtures of chiral compounds to the sample gas as reference substances and ionizing them by the same
laser pulse. Therefore, in the mass spectrum of every single laser pulse, ion signals of sample and reference species appear
both being subject to the same kind of instrumental fluctuations (in particular of laser pulse energy). In another approach,
a laser repetition rate of 200 Hz allowed averaging of large numbers of laser pulses.
相似文献
7.
Enhanced recognition of non-complementary hybridization by single-LNA-modified oligonucleotide probes 总被引:1,自引:0,他引:1
XianYu Piao Ying Yan Jing Yan YiFu Guan 《Analytical and bioanalytical chemistry》2009,394(6):1637-1643
Locked nucleic acid (LNA) is a deoxyribonucleotide analogue with an unusual ‘locked’ furanose conformation. LNA-modified oligonucleotide
probes have demonstrated an enhanced binding affinity towards their complementary strands; however, their potential to discriminate
non-complementary hybridization of mismatches has not been explored. In this study, we investigated the effect of the chemical
nature of LNA nucleobases on the hybridization stability and the capability of LNA-modified oligonucleotides to discriminate
the LNA:DNA mismatched base pairs. It was observed that LNA modification indeed improves the discrimination capability of
oligonucleotides by increasing the melting temperature differences between the complementary duplexes and hybrids containing
mismatches. Particularly, LNA purines offer a greater potential to recognize the mismatches than LNA pyrimidines and DNA purines.
Real-time PCR experiments further confirmed that LNA modifications at the 3′-end are more effective. The results and conclusions
in this study provide useful information for hybridization-based nucleic acid analysis where designing sound oligonucleotide
probes is crucial to the success of the analyses.
Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. 相似文献
8.
Kurt Kalcher Ivan Svancara Marijo Buzuk Karel Vytras Alain Walcarius 《Monatshefte für Chemie / Chemical Monthly》2009,140(8):861-889
Abstract An overview of the use of electrochemical sensors made from heterogeneous carbon materials (carbon paste electrodes, screen-printed
carbon electrodes) in the field of food analysis is presented. Sensors for inorganic and organic analytes as well as biosensors
are summarized.
Graphical abstract
相似文献
9.
Emily O’Neill Danielle Harrington John Allison 《Analytical and bioanalytical chemistry》2009,393(8):2029-2038
Monitoring of cell cultures in microbioreactors is a crucial task in cell bioassays and toxicological tests. In this work
a novel tool based on a miniaturized sensor array fabricated using low-temperature cofired ceramics (LTCC) technology is presented.
The developed device is applied to the monitoring of cell-culture media change, detection of the growth of various species,
and in toxicological studies performed with the use of cells. Noninvasive monitoring performed with the LTCC microelectrode
array can be applied for future cell-engineering purposes.
Figure Microelectrode array for monitoring of cell cultures 相似文献
10.
Kuroda D Zhang Y Wang J Kaji N Tokeshi M Baba Y 《Analytical and bioanalytical chemistry》2008,391(7):2543-2549
A thermo-responsive separation matrix, consisting of Pluronic F127 tri-block copolymers of poly(ethylene oxide) and poly(propylene
oxide), was used to separate DNA fragments by microchip electrophoresis. At low temperature, the polymer matrix was low in
viscosity and allowed rapid loading into a microchannel under low pressure. With increasing temperatures above 25°C, the Pluronic
F127 solution forms a liquid crystalline phase consisting of spherical micelles with diameters of 17–19 nm. The solution can
be used to separate DNA fragments from 100 bp to 1500 bp on poly(methyl methacrylate) (PMMA) chips. This temperature-sensitive
and viscosity-tunable polymer provided excellent resolution over a wide range of DNA sizes. Separation is based on a different
mechanism compared with conventional matrices such as methylcellulose. To illustrate the separation mechanism of DNA in a
Pluronic F127 solution, DNA molecular imaging was performed by fluorescence microscopy with F127 polymer as the separation
matrix in microchip electrophoresis.
Figure Temperature dependence of the viscosity of 20% w/w Pluronic F127 solution in 1xTBE buffer. Dotted approximates resultant curve. 相似文献
11.
Tuulia Hyötyläinen 《Analytical and bioanalytical chemistry》2009,394(3):743-758
Sample preparation before chromatographic separation is the most time-consuming and error-prone part of the analytical procedure.
Therefore, selecting and optimizing an appropriate sample preparation scheme is a key factor in the final success of the analysis,
and the judicious choice of an appropriate procedure greatly influences the reliability and accuracy of a given analysis.
The main objective of this review is to critically evaluate the applicability, disadvantages, and advantages of various sample
preparation techniques. Particular emphasis is placed on extraction techniques suitable for both liquid and solid samples.
Figure Miniaturised extraction techniques allow sensitive analysis of also small sample volumes. 相似文献
12.
Sen Hou Xinxin Li Xiaoyu Li Xi-Zeng Feng Rui Wang Chen Wang Lei Yu Ming-Qiang Qiao 《Analytical and bioanalytical chemistry》2009,394(3):783-789
Surface wettability conversion with hydrophobins is important for its applications in biodevices. In this work, the application
of a type I hydrophobin HGFI in surface wettability conversion on mica, glass, and poly(dimethylsiloxane) (PDMS) was investigated.
X-ray photoelectron spectroscopy (XPS) and water-contact-angle (WCA) measurements indicated that HGFI modification could efficiently
change the surface wettability. Data also showed that self-assembled HGFI had better stability than type II hydrophobin HFBI.
Protein patterning and the following immunoassay illustrated that surface modification with HGFI should be a feasible strategy
for biosensor device fabrication.
Figure A hydrophobin HGFI has been applied into surface wettability conversion for protein immobilization
Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. 相似文献
13.
Subash C. B. Gopinath Koichi Awazu Makoto Fujimaki Katsuaki Sugimoto Yoshimichi Ohki Tetsuro Komatsubara Junji Tominaga Penmetcha K. R. Kumar 《Analytical and bioanalytical chemistry》2009,394(2):481-488
Biological self-assembly is a natural process that involves various biomolecules, and finding the missing partner in these
interactions is crucial for a specific biological function. Previously, we showed that evanescent-field-coupled waveguide-mode
sensor in conjunction with a SiO2 waveguide, the surfaces which contain cylindrical nanometric holes produced by atomic bombardment, allowed us to detect efficiently
the biomolecular interactions. In the present studies, we showed that the assembly of biomolecules can be monitored using
the evanescent-field-coupled waveguide-mode biosensor and thus provide a methodology in monitoring assembly process in macromolecular
machines while they are assembling.
Evanescent-field-coupled waveguide-mode sensor
Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. 相似文献
14.
Aristeidis E. Niotis Christos Mastichiadis Panagiota S. Petrou Ion Christofidis Sotirios E. Kakabakos Athanasia Siafaka-Kapadai Konstantinos Misiakos 《Analytical and bioanalytical chemistry》2010,396(3):1187-1196
The early diagnosis of acute myocardial infarction requires the determination of several markers in serum shortly after its
incidence. The markers most widely employed are the isoenzyme MB of creatine kinase (CK-MB) and the cardiac troponin I (cTnI).
In the present work, a capillary waveguide fluoroimmunosensor for fast and highly sensitive simultaneous determination of
these markers in serum samples is demonstrated. The dual-analyte immunosensor was realized using glass capillaries internally
modified with an ultrathin poly(dimethylsiloxane) film by creating discrete bands of analyte-specific antibodies. The capillary
was then filled with a mixture of sample and biotinylated detection antibodies followed by reaction with streptavidin–horseradish
peroxidase and incubation with a fluorescently labeled tyramide derivative to accumulate fluorescent labels onto immunoreaction
bands. Upon scanning the capillary with a laser beam, part of the emitted fluorescence is trapped and waveguided through the
capillary wall to a photomultiplier placed on one of its ends. The employment of tyramide signal amplification provided detection
limits of 0.2 and 0.5 ng/mL for cTnI and CK-MB, respectively, in a total assay time of 30 min compared to 0.8 and 0.6 ng/mL
obtained for the corresponding assays when the conventional fluorescent label R-phycoerythrin was used in a 65-min assay.
In addition, the proposed immunosensor provided accurate and repeatable measurements (intra-assay and interassay coefficients
of variation lower than 10%), and the values determined in serum samples were in good agreement with those obtained with commercially
available enzyme immunoassays. Thus, the proposed capillary waveguide fluoroimmunosensor has all the required characteristics
for fast and reliable diagnosis of acute myocardial infarction.
相似文献
15.
András Gergely Péter Horváth György Szász Gábor Veress 《Analytical and bioanalytical chemistry》2009,394(8):2105-2109
A three-step gradient reversed-phase high-performance liquid chromatography (RP-HPLC) method was developed for the separation
of dehydroepiandrosterone (DHEA), its sulfate ester (DHEA-S), its three C7-oxidized metabolites (7αOH-DHEA, 7βOH-DHEA, 7-keto-DHEA),
and its biosynthetic congeners (androstenedione, testosterone, estradiol, pregnenolone). This new method allows the quantitative
characterization of DHEA metabolism and biosynthetic transformation under given physiological, pathological, or therapeutically
influenced circumstances. Tetrahydrofuran probably acts as a proton acceptor coadsorbent, while isopropanol behaves as a proton
donor during the separation of testosterone, estradiol, and the stereoisomers of 7-OH-DHEA.
Figure Optimized gradient RP-HPLC results in full separation of DHEA from its biosynthetic congeners and metabolites 相似文献
16.
John Benedet Donglai Lu Karel Cizek Jeff La Belle Joseph Wang 《Analytical and bioanalytical chemistry》2009,395(2):371-376
Rapid detection of the hydrogen peroxide precursor of peroxide explosives is required in numerous security screening applications.
We describe a highly sensitive and selective amperometric detection of hydrogen peroxide vapor at an agarose-coated Prussian-blue
(PB) modified thick-film carbon transducer. The sensor responds rapidly and reversibly to dynamic changes in the level of
the peroxide vapor, with no apparent carry over and with a detection limit of 6 ppbv. The remarkable selectivity of the PB-based
screen-printed electrode towards hydrogen peroxide leads to effective discrimination against common beverage samples. For
example, blind tests have demonstrated the ability to selectively and non-invasively identify concealed hydrogen peroxide
in drinking cups and bottles. The attractive performance of the new microfabricated PB-based amperometric peroxide vapor sensor
indicates great potential for addressing a wide range of security screening and surveillance applications.
Figure Experimental setup (left) with three electrode electrochemical Hydrogen Peroxide sensor hanging above container of “unknown” liquid. Schematic (right) demonstrating fundamental principles of operation of the sensor. 相似文献
17.
A novel quartz device has been designed to trap arsine and selenium hydride and subsequently to volatilize the collected analyte
and atomize it for atomic-absorption spectrometric detection. The device is actually the multiple microflame quartz-tube atomizer
(multiatomizer) with inlet arm modified to serve as the trap and to accommodate the oxygen-delivery capillary used to combust
hydrogen during the trapping step. The effect of relevant experimental conditions (trap temperature during trapping and hydrogen
flow rate and trap temperature during volatilization) on collection and volatilization efficiency was investigated. Under
the optimum conditions collection and volatilization efficiency for arsenic and selenium were 50 and 70%, respectively.
相似文献
18.
Rudolf Tuckermann Ljiljana Puskar Mahta Zavabeti Ryo Sekine Don McNaughton 《Analytical and bioanalytical chemistry》2009,394(5):1433-1441
An experimental apparatus combining Raman spectroscopy with acoustic levitation, Raman acoustic levitation spectroscopy (RALS),
is investigated in the field of physical and chemical analytics. Whereas acoustic levitation enables the contactless handling
of microsized samples, Raman spectroscopy offers the advantage of a noninvasive method without complex sample preparation.
After carrying out some systematic tests to probe the sensitivity of the technique to drop size, shape, and position, RALS
has been successfully applied in monitoring sample dilution and preconcentration, evaporation, crystallization, an acid–base
reaction, and analytes in a surface-enhanced Raman spectroscopy colloidal suspension.
Figure We have systematically investigated the analytical potential of Raman spectroscopy of samples in acoustically levitated drops. 相似文献
19.
Yveline Henchoz Bruno Bard Davy Guillarme Pierre-Alain Carrupt Jean-Luc Veuthey Sophie Martel 《Analytical and bioanalytical chemistry》2009,394(3):707-729
The measurement of physicochemical properties at an early phase of drug discovery and development is crucial to reduce attrition
rates due to poor biopharmaceutical properties. Among these properties, ionization, lipophilicity, solubility and permeability
are mandatory to predict the pharmacokinetic behavior of NCEs (new chemical entities). Due to the high number of NCEs, the
analytical tools used to measure these properties are automated and progressively adapted to high-throughput technologies.
The present review is dedicated to experimental methods applied in the early drug discovery process for the determination
of solubility, ionization constants, lipophilicity and permeability of small molecules. The principles and experimental conditions
of the different methods are described, and important enhancements in terms of throughput are highlighted.
Figure Scheme of the Drug Research Process. 相似文献
20.
Bo Xu Xiaojun Feng Youzhi Xu Wei Du Qingming Luo Bi-Feng Liu 《Analytical and bioanalytical chemistry》2009,394(7):1911-1917
Analysis of complex biological samples requires the use of high-throughput analytical tools. In this work, a microfluidic
two-dimensional electrophoresis system was developed with mercury-lamp-induced fluorescence detection. Mixtures of 20 standard
amino acids were used to evaluate the separation performance of the system. After fluorescent labeling with fluorescein isothiocyanate,
mixtures of amino acids were separated by micellar electrokinetic chromatography in the first dimension and by capillary zone
electrophoresis in the second. A double electrokinetic valve system was employed for the sample injection and the switching
between separation channels. Under the optimized conditions, 20 standard amino acids were effectively separated within 20 min
with high resolution and repeatability. Quantitative analysis revealed linear dynamic ranges of over three orders of magnitudes
with detection limits at micromolar range. To further evaluate the reliability of the system, quantitative analysis of a commercial
nutrition supplement liquid was successfully demonstrated.
Figure 相似文献