Current Analytical Chemistry - Volume 2, Issue 1, 2006

Quantification of gene expression is the sine qua non in understanding how the components of a biological cell or an organism change and interact following external or internal perturbations. Analytical techniques that combine stable isotopic protein and peptides labelling and mass spectrometric analysis are becoming increasingly popular in determining protein abundance. The common motif, which recurs in these bio-analytical methods, is the parallel labelling of the protein samples with stable isotopes. The ratio of the differently labelled peptide ions observed in the mass spectrum reflects the differences in concentration between the species in the combined mixture. Isotopes can be introduced using labelled nutrients prior to proteolytic digestion or by chemical derivatisation of the peptide functional groups after proteolysis. Although metabolic incorporation of labelled isotopes provides a more global quantification strategy, chemical derivatisation based approaches are also suitable for those laboratories with basic proteomics facilities. Advantages and limitations of these techniques are reviewed, highlighting examples of specific biological applications.

Atmospheric pressure matrix-assisted laser desorption/ionization (AP-MALDI) was first introduced in 1999. In the last few years, there has been a rapid growth in publications describing the development and application of APMALDI to the mass spectrometric analysis of peptides, polymers and oligosaccharides. This short review discusses recent advances in AP-MALDI, focusing on the analysis of biomolecules and the relative performance of MALDI at atmospheric pressure and in vacuo.

This review article deals with recent development in the analyses of proteins and DNA by capillary electrophoresis (CE) and microchip CE (MCE) using polymer solutions. Polymer solutions are commonly prepared from linear polymers such as linear polyacrylamide, poly(ethylene oxide), hydroxyethyl cellulose, and poly(vinylpyrrolidone), which possess the advantages of low fluorescence background, self-coating capability, and high sieving ability. We discuss the key parameters that affect the separation efficiency, including electroosmotic flow, chemical and physical properties of polymers, background electrolytes, and additives. The advantages and disadvantages of CE and MCE in conjunction with laser-induced fluorescence using polymer solutions for the analyses of DNA and proteins in biological fluids are highlighted. Several recently developed CE approaches such as gradient techniques and sample stacking for efficient and sensitive analyses of DNA and proteins are emphasized. We also address the trend of using nanomaterials for bioanalysis in CE and MCE.

Affinity capillary electrophoresis (ACE) is a versatile analytical technique that has been shown to be an efficient and accurate tool to probe non-covalent interactions and to determine binding and dissociation constants between receptors and ligands. ACE uses as its basis the change in migration time of a receptor upon binding to a ligand generally found in the electrophoresis buffer. Subsequent analysis using non-interacting standards realizes values for the binding constant. The technique has a number of advantages over other binding assay methods in that binding parameters can be obtained expeditiously, reproducibly, and with minimal sample quantity requirements and preparation. This review focuses on the literature describing the use of ACE from January 2003 to July 2004.

This paper reviews recent advances and the key strategies in microchip capillary electrophoresis (CE) with electrochemical detection (ECD) for separating and detecting a variety of environmental pollutants. The subjects covered include the fabrication of microfluidic chips, sample pretreatments, ECD, typical applications of microchip CE with ECD in environmental analysis, and future prospects. It is expected that microchip CE-ECD will become a powerful tool in the environmental field and will lead to the creation of truly portable devices.

The determination of superoxide anion (O2 .-) is of great physiological and pathological importance since O2 .- is considered to be involved in the etiology of aging, cancer, and progressive neurodegenerative diseases such as Parkinson's disease. The determination of the local concentration of O2 .- has been a relatively challenging analytical problem because of the low concentration, high reactivity and short lifetime of O2 .- in biological systems. This review raises a number of methods, highlighting electrochemical methodologies, for the determination of O2 .-. These electrochemical sensors and biosensors and their analytical characteristics are discussed in detail.

Semiconductor quantum dots (QDs) hold immense promise as versatile fluorescent probes in biological staining and diagnostics. Compared with their organic dyes counterparts, the ideal optical properties of QDs (e.g., high photobleaching threshold, characteristic narrow and symmetric fluorescent spectra, size-tunable emission and simultaneous excitation) offer a possibility to tag biomolecules in ultra-sensitive biological detection. These unique photophysical characteristics of QDs also promote the invention of using QD-tagged microbeads as fluorescent probes for biological applications including multiplexed bioassays, high-throughput screening and combinatorial chemistry. This mini-review describes briefly some background knowledge about QDs, the synthesis of monodispersion polymeric and silica microbeads, the preparation and some of the biological applications of QD-tagged microbeads. Other concepts related to QD-tagged microbeads are discussed.

At present, proteomics that promises deep understanding of genomic functions is developing rapidly. Proteomics at single cell level represents new challenges that greatly call for technique innovation in current analytical chemistry. In this mini-review, the concept of single cell proteomics (SCP) is clarified. Recent advancement on strategy and methodology of SCP is reviewed, with emphasis on micro-separation technique improvement such as ultra-thin layer gel electrophoresis, multidimensional micro liquid chromatography and capillary electrophoresis, and microfluidic chip as well. Future development on SCP is also suggested.

Endocrine disrupting chemicals (EDCs) affect the internal endocrine functions in human and thus effective analytical methods have been developed in recent years. In particular, quantification of EDCs in human biological fluids is essential to obtain direct information about the health risk associated with exposure to these compounds. High performance liquid chromatography (HPLC) has often been applied to the measurement of relatively polar EDCs for instance bisphenol A, phytoestrogen and phthalte metabolites in human biological samples. For detection of EDCs by HPLC, several detection methods such as ultraviolet, fluorescence, electrochemical detection and mass spectrometry are used. These detection methods are selected appropriately in consideration to their sensitivity and selectivity, and characteristics of analytes. Recently, the use of tandem mass spectrometry has increased owing to its excellent specificity. These HPLC methods are used in combination with a suitable sample preparation technique to remove large amounts of complex matrices that interfere with the detection of trace amounts of EDCs in biological fluids and to obtain sufficiently clean chromatogram. We summarized the detection and preparation method for the HPLC determination of EDCs and their metabolites in various human biological fluids such as blood, urine and breast milk.

During last years the application of chemical analysis to historical, artistic and archaeological specimens has gained more and more importance. The integration of scientific studies with information derived from stylistic or historic knowledge is really useful for the purposes of conservation, authentication, dating and attribution. Since the samples under examination are unique and irreplaceable the specimens must be submitted to non-destructive or at least microdestructive analysis. This work shows an overview of the state-of-the-art of vibrational spectroscopic techniques (mainly FTIR and Raman spectroscopies) as an analytical probe in artwork diagnosis, focusing on the need for specific spectra databases to perform accurate determinations. Some applications carried out by using both spectroscopic techniques will be presented.