Showing papers in "Analyst in 2009"
TL;DR: It is demonstrated that the NMR data as such reveal associations between systemic metabolic phenotypes and the metabolic syndrome, paving the way for this technology in large-scale clinical and epidemiological studies.
Abstract: A high-throughput proton (1H) nuclear magnetic resonance (NMR) metabonomics approach is introduced to characterise systemic metabolic phenotypes. The methodology combines two molecular windows that contain the majority of the metabolic information available by 1H NMR from native serum, e.g. serum lipids, lipoprotein subclasses as well as various low-molecular-weight metabolites. The experimentation is robotics-controlled and fully automated with a capacity of about 150–180 samples in 24 h. To the best of our knowledge, the presented set-up is unique in the sense of experimental high-throughput, cost-effectiveness, and automated multi-metabolic data analyses. As an example, we demonstrate that the NMR data as such reveal associations between systemic metabolic phenotypes and the metabolic syndrome (n = 4407). The high-throughput of up to 50 000 serum samples per year is also paving the way for this technology in large-scale clinical and epidemiological studies. In contradiction to single ‘biomarkers’, the application of this holistic NMR approach and the integrated computational methods provides a data-driven systems biology approach to biomedical research.
513 citations
TL;DR: This Critical Review discusses recent growth areas in fluorescent PET sensors by emphasizing the modular features of the 'fluorophore-spacer-receptor' design by critically examined as a case in point.
Abstract: Fluorescent sensors are an important part of the analytical scientist's toolbox. The use of fluorescent PET (Photoinduced Electron Transfer) sensors has seen particular growth in recent times. This Critical Review discusses recent growth areas in fluorescent PET sensors by emphasizing the modular features of the ‘fluorophore–spacer–receptor’ design. The occurrence of the dipicolylamine receptor in PET sensor designs is critically examined as a case in point.
456 citations
TL;DR: This review provides a critical overview of many analytical methods for the characterization of nanomaterials as well as their impacts on in vitro and in vivo function from the perspective of an analytical chemist and is intended to be used as a reference for scientists interested in conducting nanotoxicological research.
Abstract: During the past 20 years, improvements in nanoscale materials synthesis and characterization have given scientists great control over the fabrication of materials with features between 1 and 100 nm, unlocking many unique size-dependent properties and, thus, promising many new and/or improved technologies. Recent years have found the integration of such materials into commercial goods; a current estimate suggests there are over 800 nanoparticle-containing consumer products (The Project on Emerging Nanotechnologies Consumer Products Inventory, , accessed Oct. 2008), accounting for 147 billion USD in products in 2007 (Nanomaterials state of the market Q3 2008: stealth success, broad impact, Lux Research Inc., New York, NY, 2008). Despite this increase in the prevalence of engineered nanomaterials, there is little known about their potential impacts on environmental health and safety. The field of nanotoxicology has formed in response to this lack of information and resulted in a flurry of research studies. Nanotoxicology relies on many analytical methods for the characterization of nanomaterials as well as their impacts on in vitro and in vivo function. This review provides a critical overview of these techniques from the perspective of an analytical chemist, and is intended to be used as a reference for scientists interested in conducting nanotoxicological research as well as those interested in nanotoxicological assay development.
385 citations
TL;DR: In this article, boron-doped diamond (BDD) films are synthesized by chemical vapor deposition on various substrates to provide electrical conductivity, which is important for detecting and/or identifying species in solution.
Abstract: In recent years, conductive diamond electrodes for electrochemical applications have been a major focus of research and development. The impetus behind such endeavors could be attributed to their wide potential window, low background current, chemical inertness, and mechanical durability. Several analytes can be oxidized by conducting diamond compared to other carbon-based materials before the breakdown of water in aqueous electrolytes. This is important for detecting and/or identifying species in solution since oxygen and hydrogen evolution do not interfere with the analysis. Thus, conductive diamond electrodes take electrochemical detection into new areas and extend their usefulness to analytes which are not feasible with conventional electrode materials. Different types of diamond electrodes, polycrystalline, microcrystalline, nanocrystalline and ultrananocrystalline, have been synthesized and characterized. Of particular interest is the synthesis of boron-doped diamond (BDD) films by chemical vapor deposition on various substrates. In the tetrahedral diamond lattice, each carbon atom is covalently bonded to its neighbors forming an extremely robust crystalline structure. Some carbon atoms in the lattice are substituted with boron to provide electrical conductivity. Modification strategies of doped diamond electrodes with metallic nanoparticles and/or electropolymerized films are of importance to impart novel characteristics or to improve the performance of diamond electrodes. Biofunctionalization of diamond films is also feasible to foster several useful bioanalytical applications. A plethora of opportunities for nanoscale analytical devices based on conducting diamond is anticipated in the very near future
371 citations
TL;DR: In this article, the authors use synchrotron radiation FTIR micro-spectroscopy to record spectra of mono-dispersed poly(methyl methacrylate) (PMMA) spheres of systematically varying size and demonstrate that the spectral distortions in the data can be understood in terms of resonant Mie scattering.
Abstract: Infrared spectroscopic cytology is potentially a powerful clinical tool. However, in order for it to be successful, practitioners must be able to extract reliably a pure absorption spectrum from a measured spectrum that often contains many confounding factors. The most intractable problem to date is the, so called, dispersion artefact which most prominently manifests itself as a sharp decrease in absorbance on the high wavenumber side of the amide I band in the measured spectrum, exhibiting a derivative-like line shape. In this paper we use synchrotron radiation FTIR micro-spectroscopy to record spectra of mono-dispersed poly(methyl methacrylate) (PMMA) spheres of systematically varying size and demonstrate that the spectral distortions in the data can be understood in terms of resonant Mie scattering. A full understanding of this effect will enable us to develop strategies for deconvolving the scattering contribution and recovering the pure absorption spectrum, thus removing one of the last technological barriers to the development of clinical spectroscopic cytology.
293 citations
TL;DR: This review summarises recent advances in vibrational spectroscopy and their impact on the diagnosis of cancer and illustrates the complementary advantage of using Raman and FTIR spectroscopic technologies together.
Abstract: Vibrational spectroscopy techniques have demonstrated potential to provide non-destructive, rapid, clinically relevant diagnostic information. Early detection is the most important factor in the prevention of cancer. Raman and infrared spectroscopy enable the biochemical signatures from biological tissues to be extracted and analysed. In conjunction with advanced chemometrics such measurements can contribute to the diagnostic assessment of biological material. This paper also illustrates the complementary advantage of using Raman and FTIR spectroscopy technologies together. Clinical requirements are increasingly met by technological developments which show promise to become a clinical reality. This review summarises recent advances in vibrational spectroscopy and their impact on the diagnosis of cancer.
278 citations
TL;DR: The combination of accurate mass data for a large collection of metabolites, theoretical isotope abundance data and knowledge of the different ion types detected provided a greater number of electrospray mass spectrometric signals which were putatively identified and with greater confidence in the samples studied.
Abstract: The chemical identification of mass spectrometric signals in metabolomic applications is important to provide conversion of analytical data to biological knowledge about metabolic pathways. The complexity of electrospray mass spectrometric data acquired from a range of samples (serum, urine, yeast intracellular extracts, yeast metabolic footprints, placental tissue metabolic footprints) has been investigated and has defined the frequency of different ion types routinely detected. Although some ion types were expected (protonated and deprotonated peaks, isotope peaks, multiply charged peaks) others were not expected (sodium formate adduct ions). In parallel, the Manchester Metabolomics Database (MMD) has been constructed with data from genome scale metabolic reconstructions, HMDB, KEGG, Lipid Maps, BioCyc and DrugBank to provide knowledge on 42,687 endogenous and exogenous metabolite species. The combination of accurate mass data for a large collection of metabolites, theoretical isotope abundance data and knowledge of the different ion types detected provided a greater number of electrospray mass spectrometric signals which were putatively identified and with greater confidence in the samples studied. To provide definitive identification metabolite-specific mass spectral libraries for UPLC-MS and GC-MS have been constructed for 1,065 commercially available authentic standards. The MMD data are available at http://dbkgroup.org/MMD/.
263 citations
TL;DR: Highlights of advances in instrumentation and applications within the last three years are provided to illustrate the impact of this rapidly growing area of research.
Abstract: Isotope Ratio Mass Spectrometry (IRMS) is a specialized technique used to provide information about the geographic, chemical, and biological origins of substances. The ability to determine the source of an organic substance stems from the relative isotopic abundances of the elements which comprise the material. Because the isotope ratios of elements such as carbon, hydrogen, oxygen, sulfur, and nitrogen can become locally enriched or depleted through a variety of kinetic and thermodynamic factors, measurement of the isotope ratios can be used to differentiate between samples which otherwise share identical chemical compositions. Several sample introduction methods are now available for commercial isotope ratio mass spectrometers. Combustion is most commonly used for bulk isotopic analysis, whereas gas and liquid chromatography are predominately used for the real-time isotopic analysis of specific compounds within a mixture. Here, highlights of advances in instrumentation and applications within the last three years are provided to illustrate the impact of this rapidly growing area of research. Some prominent new applications include authenticating organic food produce, ascertaining whether or not African elephants are guilty of night-time raids on farmers' crops, and linking forensic drug and soil samples from a crime scene to a suspected point of origin. For the sake of brevity, we focus this Minireview on the isotope ratio measurements of lighter-elements common to organic sources; we do not cover the equally important field of inorganic isotope ratio mass spectrometry.
245 citations
TL;DR: Applications of Raman spectroscopy to cells and tissues that are expected to gain significance in the future such as the combination with imaging approaches, microscopy, optical traps and fiber-optic probes are summarized.
Abstract: Raman spectroscopy has been recognized to be a powerful tool to study cells and tissues because the method provides molecular information without external markers such as stains or radioactive labels. To overcome the disadvantage of low signal intensities from most biomolecules, enhancement effects are utilized. A non-linear variant of Raman spectroscopy called coherent anti-Stokes Raman spectroscopy (CARS) belongs to the most promising techniques because it combines signal enhancement due to the coherent nature of the process with further advantages such as directional emission, narrow spectral bandwidth and no disturbing interference with autofluorescence. This review describes briefly the principles of the methods and summarizes applications to cells and tissues that are expected to gain significance in the future such as the combination with imaging approaches, microscopy, optical traps and fiber-optic probes.
243 citations
TL;DR: Advances in sensitivity and instrumentation now allow CFMEs to be used for measurements of natural dopamine release that occur during behavioral experiments, and a new technique combining electrochemistry with electrophysiology at a single microelectrode facilitates a better understanding of neurotransmitter concentrations and their effects on cell firing.
Abstract: Carbon-fiber microelectrodes (CFMEs) have been a useful tool for measuring rapid changes in neurotransmitters because of their small size, sensitivity, and good electrochemical properties. In this article, we highlight recent advances using CFMEs for measuring neurotransmitters in vivo. Dopamine has been a primary neurotransmitter of interest but direct electrochemical detection of other neurochemicals including nitric oxide and adenosine has also been investigated. Surface treatments have been studied to enhance electrode sensitivity, such as covalent modification or the addition of a layer of carbon nanotubes. Enzyme-modified microelectrodes that detect non-electroactive compounds further extend the usefulness of CFMEs beyond the traditional monoamines. CFMEs continue to be used in vivo to understand basic neurobiological mechanisms and the actions of pharmacological agents, including drugs of abuse. Advances in sensitivity and instrumentation now allow CFMEs to be used for measurements of natural dopamine release that occur during behavioral experiments. A new technique combining electrochemistry with electrophysiology at a single microelectrode facilitates a better understanding of neurotransmitter concentrations and their effects on cell firing. Future research in this field will likely concentrate on fabricating smaller electrodes and electrode arrays, as well as expanding the use of CFMEs in neuroscience beyond dopamine.
191 citations
TL;DR: The robust nature of oligonucleotides, in terms of synthesis, storage, and wide range of temperature stability and chemical manipulation, makes them highly suitable for biosensor design and engineering.
Abstract: Oligonucleotides were once considered only functional as molecules for the storage of genetic information. However, the discovery of RNAzymes, and later, DNAzymes, unravelled the innate potential of oligonucleotides in many other biological applications. In the last two decades, these applications have been further expanded through the introduction of Systematic Evolution of Ligands by EXponential enrichment (SELEX) which has generated, by repeated rounds of in vitro selection, a type of molecular probe termed aptamers. Aptamers are oligonucleic acid (or peptide) molecules that can bind to various molecular targets and are viewed as complements to antibodies. Aptamers have found applications in many areas, such as bio-technology, medicine, pharmacology, microbiology, and analytical chemistry, including chromatographic separation and biosensors. In this review, we focus on the use of aptamers in the development of biosensors. Coupled with their ability to bind a variety of targets, the robust nature of oligonucleotides, in terms of synthesis, storage, and wide range of temperature stability and chemical manipulation, makes them highly suitable for biosensor design and engineering. Among the many design strategies, we discuss three general paradigms that have appeared most frequently in the literature: structure-switching, enzyme-based, and aptazyme-based designs.
TL;DR: A low temperature plasma probe constructed using dielectric barrier discharge (DBD) and employed for the detection of explosives on a variety of substrates under ambient conditions allowed direct detection of trace explosives on both conductive and non-conductive substrates, thus expanding the applicability ofLow temperature plasma desorption mass spectrometry.
Abstract: In this paper, we have constructed a low temperature plasma (LTP) probe using dielectric barrier discharge (DBD) and employed it for the detection of explosives on a variety of substrates under ambient conditions. Upon discharge, a transient, low-temperature non-equilibrium plasma comprising ions, electrons and metastable atoms are generated between the electrodes. Three common explosives, 2,4,6-trinitrotoluene (TNT), hexahydro-1,3,5-trinitro-1,3,5-trinitro-1,3,5-triazine (RDX), and pentaerythritol tetranitrate (PETN), were directly desorbed and ionized from solid surfaces, followed by subsequent analysis using the mass spectrometer in the negative ion mode. Limits of detection (LODs) were 500 fg for TNT, 1 pg for RDX, and 500 fg for PETN. The reliability of the method was characterized by a successful analysis of a mixture of the three explosives. The ion source also allowed direct detection of trace explosives on both conductive and non-conductive substrates, thus expanding the applicability of low temperature plasma desorption mass spectrometry.
TL;DR: New aptamer probes for non-small cell lung cancer (NSCLC) are developed by directing the aptamer selection process against the living cells of adenocarcinoma, the most common subtype of NSCLC by generating and evaluating single-stranded DNA aptamers.
Abstract: In this work, we have developed new aptamer probes for non-small cell lung cancer (NSCLC) by directing the aptamer selection process against the living cells of adenocarcinoma, the most common subtype of NSCLC. A panel of single-stranded DNA (ssDNA) aptamers were generated and evaluated for adenocarcinoma cell recognition. The aptamers bound to the adenocarcinoma cells with dissociation constants in the nanomolar range and the binding of the selected aptamers to the adenocarcinoma cells were significantly stronger than the other cancerous lung cells as well as other types of cancer cells. Moreover, the application of the aptamers to the clinical tissue section samples showed the differentiation of adenocarcinoma from normal lung tissue and other subtypes of lung cancer. The aptamers are expected to be new molecular probes for the investigation of the molecular bases of different NSCLC subtypes and their biological heterogeneity, which is valuable for advancing NSCLC diagnosis and treatment.
TL;DR: Spectrum-wide relative standard deviations are calculated for ten metabolomics datasets, spanning a variety of sample types from mammals, fish, invertebrates and a cell line, and displayed succinctly as boxplots to propose spectral RSDs and their median values contained herein as practical benchmarks for metabolomics studies.
Abstract: Metabolomics datasets, by definition, comprise of measurements of large numbers of metabolites. Both technical (analytical) and biological factors will induce variation within these measurements that is not consistent across all metabolites. Consequently, criteria are required to assess the reproducibility of metabolomics datasets that are derived from all the detected metabolites. Here we calculate spectrum-wide relative standard deviations (RSDs; also termed coefficient of variation, CV) for ten metabolomics datasets, spanning a variety of sample types from mammals, fish, invertebrates and a cell line, and display them succinctly as boxplots. We demonstrate multiple applications of spectral RSDs for characterising technical as well as inter-individual biological variation: for optimising metabolite extractions, comparing analytical techniques, investigating matrix effects, and comparing biofluids and tissue extracts from single and multiple species for optimising experimental design. Technical variation within metabolomics datasets, recorded using one- and two-dimensional NMR and mass spectrometry, ranges from 1.6 to 20.6% (reported as the median spectral RSD). Inter-individual biological variation is typically larger, ranging from as low as 7.2% for tissue extracts from laboratory-housed rats to 58.4% for fish plasma. In addition, for some of the datasets we confirm that the spectral RSD values are largely invariant across different spectral processing methods, such as baseline correction, normalisation and binning resolution. In conclusion, we propose spectral RSDs and their median values contained herein as practical benchmarks for metabolomics studies.
TL;DR: This review will discuss the chemical and physical properties of nanomaterials necessary for use as chemosensors and biosensors, and highlight some noteworthy recent avenues using nanoscale materials as scaffolds for chemosensing and biosensing.
Abstract: Over the past few decades the fabrication of nanoscale materials for use in chemical sensing, biomedical and biological analyses has proven a promising avenue. Nanomaterials show promise in such chemical and biological analysis mainly due to their highly tunable size- and shape-dependent chemical and physical properties. Furthermore, they exhibit unique surface chemistry, thermal stability, high surface area and large pore volume per unit mass that can be exploited for sensor fabrication. This review will discuss the chemical and physical properties of nanomaterials necessary for use as chemosensors and biosensors. It will also highlight some noteworthy recent avenues using nanoscale materials as scaffolds for chemosensing and biosensing. Nanomaterials that have proven to be useful for the fabrication of sensors, as reviewed herein, have compositions including metals, metal oxides, chalcogenides and polymers. Their structures range from nanoparticles, nanorods, and nanowires to nanoporous and core-shells. Examples of the different types of structures and compositions as well as sensors and biosensors fabricated from them will be described. Some nanomaterials are functionalized with various kinds of ligands and bioactive groups to produce sensitive and selective sensors for specific analytes. The combination of two or more types of nanostructures with core-shell type nanoassemblies and other composite structures, in addition to advantageous features enhancing sensitivity and response time of related sensors, are also discussed.
TL;DR: The present fluorescent sensor possesses remarkable selectivity for cyanide over other anions, and negligible influences were observed on the cyanide detection by the coexistence of other anion or biological species.
Abstract: A new fluorescent sensor for the sensitive and selective detection of cyanide (CN(-)) in aqueous media was developed herein. The sensing approach is based on CN(-)-modulated quenching behavior of Cu(2+) toward the photoluminescence (PL) of CdTe quantum dots (QDs). In the presence of CN(-), the PL of QDs that have been quenched by Cu(2+) was found to be efficiently recovered, which then allows the detection of CN(-) in a very simple approach. Experimental results showed that the pH of the buffer solution, concentration of copper ions, and size of CdTe QDs all influenced the response of the sensor to CN(-). Under the optimal conditions, a good linear relationship between the PL intensity and the concentration of CN(-) can be obtained in the range of 3.0 x 10(-7) to 1.2 x 10(-5) M, with a detection limit as low as 1.5 x 10(-7) M. In addition, the present fluorescent sensor possesses remarkable selectivity for cyanide over other anions, and negligible influences were observed on the cyanide detection by the coexistence of other anions or biological species (such as albumin and typical blood constituents). Therefore, we expect the proposed copper ion-modified QDs to be an efficient and reliable sensing system to monitor cyanide concentration in environmental or clinical applications.
TL;DR: Recent studies on nanodiamonds and their use as biolabels are reviewed, while highlighting the links between the physical, chemical and biological issues that arise.
Abstract: Fluorescent defects in non-cytotoxic diamond nanoparticles have recently emerged as a preferred candidate for optical labels in biological and medical imaging. The bright fluorescence at 550–800 nm originates from point defects within the particles, some of which appear naturally, while others can be artificially incorporated during synthesis or can be introduced using high-energy ion beam irradiation and subsequent thermal annealing. However, in order for the fluorescent defects to be useful in bio-medical applications there are a number of materials challenges that must be overcome. In this paper, recent studies on nanodiamonds and their use as biolabels are reviewed, while highlighting the links between the physical, chemical and biological issues that arise.
TL;DR: The designed probe exhibits high sensitivity, good photostability, excellent cell membrane permeability and strong pH dependence, and the fluorescent imaging of HepG2 cells demonstrates that the designed probe has great value in monitoring intracellular H(+) within living cells.
Abstract: A novel acidic fluorescent probe 1 has been designed, synthesized, characterized and evaluated in vivo as optical imaging of intracellular H+ The design strategy for the probe is based on the change in structure between spirocyclic (non-fluorescent) and ring-open (fluorescent) forms of rhodamine dyes The probe exhibits high sensitivity, good photostability, excellent cell membrane permeability and strong pH dependence The pH titration indicates that the fluorescence intensity increases more than 100-fold within the pH range of 42–60 with the pKa value of 485, which is valuable for studying acidic organelles in living cells The fluorescent imaging of HepG2 cells also demonstrates that the designed probe has great value in monitoring intracellular H+ within living cells
TL;DR: A novel boronate affinity monolith, poly(3-acrylamidophenylboronic acid-co-ethylene dimethacrylate) (AAPBA- co-EDMA), was prepared in 530 microm capillaries by a one-step in situ polymerization procedure using a pre-polymerization mixture.
Abstract: A novel boronate affinity monolith, poly(3-acrylamidophenylboronic acid-co-ethylene dimethacrylate) (AAPBA-co-EDMA), was prepared in 530 µm capillaries by a one-step in situpolymerization procedure using a pre-polymerization mixture consisting of functional monomer 3-acrylamidophenylboronic acid, cross-linker ethylene dimethacrylate, porogenic solvent methanol with added poly(ethylene glycol) 20 000 (PEG 20 000) and initiator azobisisobutyronitrile (AIBN). The preparation of the monolith was optimized by investigating the ratio of functional monomer to cross-linker and the effect of poly(ethylene glycol) molecular weight. The resulting boronate monolith was used as a sorbent for polymer monolith microextraction (PMME). Using nucleosides as the testing analyte, the extraction performance of this boronate monolith towards glycol-containing compounds was examined. Finally, the boronate monolith was applied for selective enrichment of glycopeptides and glycoproteins.
TL;DR: With this method, adenosine can be selectively detected in the low micromolar range, which means that the strategy reported here can be applicable to the detection of several other small target molecules.
Abstract: Based on gold nanoparticles (AuNPs) and engineered DNA aptamers, we designed a novel bioassay strategy for the detection of adenosine as a small target molecule. In this design, an aptamer is engineered to consist of two pieces of random-coil like ssDNA which are respectively attached to AuNPs through their 5′-thiol-modified end. They can reassemble into the intact aptamer tertiary structure and induce nanoparticle aggregation in the presence of the specific target. Results have demonstrated that gold nanoparticles can effectively differentiate these two different DNA structures via their characteristic surface plasmon resonance-based color change. With this method, adenosine can be selectively detected in the low micromolar range, which means that the strategy reported here can be applicable to the detection of several other small target molecules.
TL;DR: This method exhibits excellent selectivity for cysteine over other alpha-amino acids, glutathione, thioglycolic acid and mercaptoethyl alcohol.
Abstract: We report herein the development of a highly sensitive and selective colorimetric detection method for cysteine using gold nanoparticles probes. This assay relies upon the distance-dependent optical properties of gold nanoparticles, the self-assembly of cysteine on gold nanoparticles, and the interaction of a 2:1 cysteine/Cu2+ complex. In the presence of Cu2+, cysteine could rapidly induce the aggregation of gold nanoparticles, thereby resulting in red-to-blue (or purple) color change. The concentration of cysteine can be determined by monitoring with the naked eye or a UV-vis spectrometer. The present limit of detection for cysteine is 10 nM. This method exhibits excellent selectivity for cysteine over other α-amino acids, glutathione, thioglycolic acid and mercaptoethyl alcohol.
TL;DR: An assay based on the modulation in FRET efficiency between QDs and AuNPs in the presence of Pb2+, which inhibits the interaction of the QD-AuNP assembly is proposed and the detection limit is found to be 30 ppb of P b2+ due to the superior fluorescence properties of QDs.
Abstract: Positively charged CdTe-QDs capped with cysteamine (CA-CdTe-QDs) and negatively charged AuNPs capped with 11-mercaptoundecanoic acid (MUA-AuNPs) have been prepared. They are water-soluble and biocompatible. An assay for the determination of Pb2+ has been proposed based on the modulation in FRET efficiency between QDs and AuNPs in the presence of Pb2+, which inhibits the interaction of the QD-AuNP assembly. This method is easy to operate and with remarkably high sensitivity. Under the optimum conditions, the response is linearly proportional to the concentration of Pb2+ in the range 0.22–4.51 ppm, and the detection limit is found to be 30 ppb of Pb2+ due to the superior fluorescence properties of QDs. The mechanism of this strategy is also discussed.
TL;DR: These studies demonstrate the advantages of integrating metabolic correlation networks with metabonomics for finding significant potential biomarkers and can even provide biochemical insights into changes in breast cancer.
Abstract: A metabonomics strategy based on rapid resolution liquid chromatography/tandem mass spectrometry (RRLC-MS/MS), multivariate statistics and metabolic correlation networks has been implemented to find biologically significant metabolite biomarkers in breast cancer. RRLC-MS/MS analysis by electrospray ionization (ESI) in both positive and negative ion modes was employed to investigate human urine samples. The resulting data matrices were analyzed using multivariate analysis. Application of orthogonal projections to latent structures discriminate analysis (OPLS-DA) allowed us to extract several discriminated metabolites reflecting metabolic characteristics between healthy volunteers and breast cancer patients. Correlation network analysis between these metabolites has been further applied to select more reliable biomarkers. Finally, high resolution MS and MS/MS analyses were performed for the identification of the metabolites of interest. We identified 12 metabolites as potential biomarkers including amino acids, organic acids, and nucleosides. They revealed elevated tryptophan and nucleoside metabolism as well as protein degradation in breast cancer patients. These studies demonstrate the advantages of integrating metabolic correlation networks with metabonomics for finding significant potential biomarkers: this strategy not only helps identify potential biomarkers, it also further confirms these biomarkers and can even provide biochemical insights into changes in breast cancer.
TL;DR: An automated method for the removal of broad and/or moderately featured background signal is described and has been applied to spectra from biofilms of Streptococcus mutans.
Abstract: The use of Raman spectroscopy for biomedical applications requires overcoming the obstacle of the broad background that is also generated by biological samples. This background, which is often largely attributed to fluorescence, is frequently orders of magnitude greater than the Raman signal and needs to be removed in order to use Raman spectra in sample analysis. Several methods have been proposed for removing fluorescent signal, both instrumental and computational. Of the computational methods, polynomial fitting has become increasingly popular. Typically, a polynomial of approximately fifth order is used in the fitting. This method alone is not always capable of fitting some more tightly featured spectra that may be present in data, potentially coming from a contaminant in the sample itself or from the experimental design. If this signal is present in varying amounts, the polynomial background removal method can leave the residual spectra with non-uniform artifacts that hinder classification results. If a reference spectrum can be obtained for this interfering signal, however, it can be incorporated into the polynomial fit and removed separately. An automated method for the removal of broad and/or moderately featured background signal is described. In addition to simulations, the method has been applied to spectra from biofilms of Streptococcus mutans.
TL;DR: It is demonstrated that the spectrum at any point is a weighted sum of the sample reflection and transmission and that the dominance of the reflection spectrum in optically dense regions can account for some of the spectral distortions previously attributed to dispersion artefacts.
Abstract: Fourier transform infrared spectra of a single cell in transflection geometry are seen to vary significantly with position on the cell, showing a distorted derivative-like lineshape in the region of the optically dense nucleus. A similar behaviour is observable in a model system of the protein albumin doped in a potassium bromide disk. It is demonstrated that the spectrum at any point is a weighted sum of the sample reflection and transmission and that the dominance of the reflection spectrum in optically dense regions can account for some of the spectral distortions previously attributed to dispersion artefacts. Rather than being an artefact, the reflection contribution is ever present in transflection spectra and it is further demonstrated that the reflection characteristics can be used for cellular mapping.
TL;DR: An optimized sample preparation method has been developed for NMR-based urinary metabonomic studies and it is suggested that the chemical shifts for the proton signals of citrate and aromatic signals of histidine be corrected prior to multivariate data analysis especially when high resolution data were employed.
Abstract: NMR-based metabonomics has been widely employed to understand the stressor-induced perturbations to mammalian metabolism. However, inter-sample chemical shift variations for metabolites remain an outstanding problem for effective data mining. In this work, we systematically investigated the effects of pH and ionic strength on the chemical shifts for a mixture of 9 urinary metabolites. We found that the chemical shifts were decreased with the rise of pH but increased with the increase of ionic strength, which probably resulted from the pH- and ionic strength-induced alteration to the ionization equilibrium for the function groups. We also found that the chemical shift variations for most metabolites were reduced to less than 0.004 ppm when the pH was 7.1-7.7 and the salt concentration was less than 0.15 M. Based on subsequent optimization to minimize chemical shift variation, sample dilution and maximize the signal-to-noise ratio, we proposed a new buffer system consisting of K(2)HPO(4) and NaH(2)PO(4) (pH 7.4, 1.5 M) with buffer-urine volume ratio of 1 : 10 for human urinary metabonomic studies; we suggest that the chemical shifts for the proton signals of citrate and aromatic signals of histidine be corrected prior to multivariate data analysis especially when high resolution data were employed. Based on these, an optimized sample preparation method has been developed for NMR-based urinary metabonomic studies.
TL;DR: The self-assembly of labeled aptamer sub-units in the presence of their substrates provides a method for the optical or electrochemical detection of the substrate and can be applied to other substrates, e.g. adenosine triphosphate.
Abstract: The self-assembly of labeled aptamer sub-units in the presence of their substrates provides a method for the optical (fluorescence) or electrochemical detection of the substrate. One of the sub-units is linked to CdSe/ZnS quantum dots (QDs), and the self-assembly of the dye-functionalized second sub-unit with the modified QDs, in the presence of cocaine, stimulates fluorescence resonance energy transfer (FRET). This enables the detection of cocaine with a detection limit corresponding to 1 × 10−6 M. Alternatively, the aptamer fragments are modified with pyrene units. The formation of a supramolecular aptamer-substrate complex allosterically stabilizes the formation of excimer supramolecular structure, and its characteristic emission is observed. In addition, the thiolated aptamer sub-unit is assembled on an Au electrode. The Methylene Blue-labeled sub-unit binds to the surface-confined fragment in the presence of cocaine. The amperometric response of the system allows the detection of cocaine with a detection limit of 1 × 10−5 M. The approach is generic and can be applied to other substrates, e.g.adenosine triphosphate.
TL;DR: The analysis of Stevia leaves has been demonstrated without any sample preparation using desorption electrospray ionization (DESI) mass spectrometry, and the presence of characteristic glycosides in an all-natural commercial Stevia dietary supplement was confirmed.
Abstract: The analysis of Stevia leaves has been demonstrated without any sample preparation using desorption electrospray ionization (DESI) mass spectrometry. Direct rapid analysis was achieved using minimal amounts of sample (∼0.15 cm × 0.15 cm leaf fragment). Characteristic constituents of the Stevia plant are observed in both the positive and negative ion modes including a series of diterpene ‘sweet’ glycosides. The presence of the glycosides was confirmed via tandem mass spectrometry analysis using collision-induced dissociation and further supported by exact mass measurements using an LTQ-Orbitrap. The analysis of both untreated and hexane-extracted dry leaves proved that DESI can be successfully used to analyze untreated leaf fragments as identical profiles were obtained from both types of samples. Characterization and semi-quantitative determination of the glycosides was achieved based on the glycoside profile within the full mass spectrum. In addition, the presence of characteristic glycosides in an all-natural commercial Stevia dietary supplement was confirmed. This study provides an example of the application of DESI to direct screening of plant materials, in this case diterpene glycosides.
TL;DR: The molecular beacon mercury(II) ion sensor was amenable to regeneration by simply unfolding the ferrocene-labeled DNA in 10 microM cysteine, and could be regenerated with no loss in signal gain upon subsequent mercury( II) ion binding.
Abstract: We have developed a selective, sensitive, and re-usable electrochemical sensor for Hg2+ ion detection. This sensor is based on the Hg2+-induced conformational change of a single-stranded DNA (ssDNA) which involves an electroactive, ferrocene-labeled DNA hairpin structure and provides strategically the selective binding of a thymine–thymine mismatch for the Hg2+ ion. The ferrocene-labeled DNA is self-assembled through S–Au bonding on a polycrystalline gold electrode surface and the surface blocked with 3-mercapto-1-propanol to form a mixed monolayer. The modified electrode showed a voltammetric signal due to a one-step redox reaction of the surface-confined ferrocenyl moiety. The ‘signal-on’ upon mercury binding could be attributed to a change in the conformation of ferrocene-labeled DNA from an open structure to a restricted hairpin structure. The differential pulse voltammetry (DPV) of the modified electrode showed a linear response of the ferrocene oxidation signal with increase of Hg2+ concentration in the range between 0.1 and 2 µM with a detection limit of 0.1 µM. The molecular beacon mercury(II) ion sensor was amenable to regeneration by simply unfolding the ferrocene-labeled DNA in 10 µM cysteine, and could be regenerated with no loss in signal gain upon subsequent mercury(II) ion binding.
TL;DR: Different optical substrates are tested that would best preserve cell integrity and allow direct measurement of Raman spectra at the single living cell level and quartz was the most appropriate taking into consideration both cell morphology and proliferation rate.
Abstract: Raman microspectroscopy allows probing subcellular compartments and provides a unique spectral fingerprint indicative of endogenous molecular composition. Although several spectroscopic cell studies have been reported on fixed samples, only few attempts concern single growing cells. Here, we have tested different optical substrates that would best preserve cell integrity and allow direct measurement of Raman spectra at the single living cell level. Calu-1 lung cancer cells were used as a model and their morphology and growth were assessed on Raman substrates like quartz, calcium fluoride, and zinc selenide. Data show that quartz was the most appropriate taking into consideration both cell morphology and proliferation rate (47% on quartzvs. 55% of BrdU-positive cells on conventional plastic). Using quartz, 40 cells were analysed and Raman spectra were collected from nuclei and cytoplasms using a 785 nm laser excitation of 30 mW at the sample, in the spectral range of 580–1750 cm−1, and an acquisition time of 2 × 10 sec/spectrum. Discriminant spectral information related to nucleus and cytoplasm were extracted by multivariate statistical methods and attributed to nucleic acids, lipids, and proteins. Finally, Raman spectral imaging was performed to show the distribution of these components within the cell.