3 Biochemical and clinical analysis

Abstracts

3 Biochemical and clinical analysis     

3 Biochemical and clinical analysis

Abstracts

3 Biochemical and clinical analysis     

3 Biochemical and clinical analysis

Abstracts

3 Biochemical and clinical analysis     

3 Biochemical and clinical analysis

Abstracts

3 Biochemical and clinical analysis     

3 Biochemical and clinical analysis

Abstracts

     

3 Biochemical and clinical analysis

Abstracts

     

3 Biochemical and clinical analysis

Abstracts

3 Biochemical and clinical analysis     

Biochemical and clinical analysis

Abstracts

Biochemical and clinical analysis     

Biochemical and clinical analysis

Abstracts

Biochemical and clinical analysis     

Biochemical and clinical analysis by enthalpimetric measurements — a realistic alternative approach?

The contribution of enthalpimetric measurements to biochemical and clinical analysis is evaluated. After a brief discussion of instrumental considerations, non-enzymatic and enzymatic approaches to biochemical and clinical enthalpimetry are considered. The main focus of the review is on the incorporation of enzyme-catalyzed reactions with enthalpimetric measurements to provide analytical selectivity; determinations of enzyme activity, substrate concentration, inhibitor concentration and biochemical constants are discussed. Enthalpimetric devices based on immobilized enzyme technology, e.g. flow enthalpimeters and enzyme-coated thermistors and the latest technique to evolve, thermometric enzyme-linked immunoassay are treated separately. The advantages and limitations of enthalpimetry in these areas of analysis are discussed in the context of routine clinical analysis.     

Biochemical analysis with microfluidic systems

Microfluidic systems are capillary networks of varying complexity fabricated originally in silicon, but nowadays in glass and polymeric substrates. Flow of liquid is mainly controlled by use of electroosmotic effects, i.e. application of electric fields, in addition to pressurized flow, i.e. application of pressure or vacuum. Because electroosmotic flow rates depend on the charge densities on the walls of capillaries, they are influenced by substrate material, fabrication processes, surface pretreatment procedures, and buffer additives. Microfluidic systems combine the properties of capillary electrophoretic systems and flow-through analytical systems, and thus biochemical analytical assays have been developed utilizing and integrating both aspects. Proteins, peptides, and nucleic acids can be separated because of their different electrophoretic mobility; detection is achieved with fluorescence detectors. For protein analysis, in particular, interfaces between microfluidic chips and mass spectrometers were developed. Further levels of integration of required sample-treatment steps were achieved by integration of protein digestion by immobilized trypsin and amplification of nucleic acids by the polymerase chain reaction. Kinetic constants of enzyme reactions were determined by adjusting different degrees of dilution of enzyme substrates or inhibitors within a single chip utilizing mainly the properties of controlled dosing and mixing liquids within a chip. For analysis of kinase reactions, however, a combination of a reaction step (enzyme with substrate and inhibitor) and a separation step (enzyme substrate and reaction product) was required. Microfluidic chips also enable separation of analytes from sample matrix constituents, which can interfere with quantitative determination, if they have different electrophoretic mobilities. In addition to analysis of nucleic acids and enzymes, immunoassays are the third group of analytical assays performed in microfluidic chips. They utilize either affinity capillary electrophoresis as a homogeneous assay format, or immobilized antigens or antibodies in heterogeneous assays with serial supply of reagents and washing solutions.     

Biochemical speciation analysis by hyphenated techniques

The elucidation of mechanisms that govern the essentiality and toxicity of trace elements in living organisms is critically dependent upon the possibility of the identification, characterization and determination of chemical forms of these elements involved in life processes. The recent progress and the state-of-the-art of biochemical species-selective trace element analysis are critically evaluated with particular emphasis on the use of techniques combining the high selectivity of high performance liquid chromatography (HPLC) with the elemental or molecular specificity of mass spectrometry [using inductively coupled plasma (ICP) or electrospray ionization (ESI)]. The potential and limitations of hyphenated techniques as a tool for speciation of metals and metalloids in biological materials is discussed using a number of examples drawn from the latest research in the authors’ laboratory.     

Biochemical analysis of wood and wood products by pyrolysis-mass spectrometry and multivariate analysis

Pyrolysis—mass spectrometry in combination with discriminant analysis and a newly development non-supervised mixture analysis approach is shown to provide valuable information about the biochemical composition of wood and wood products. The application of this technique to the chemotaxonomy of sagebrush (Artemisia) species as well as to the quality control of wood pulping processes is discussed. With regard to the latter application strong correlations are found between classical lignin and xylan determinations and direct pyrolysis—mass spectrometric analysis results.     

Biochemical analysis with the expanded genetic lexicon

The information used to build proteins is stored in the genetic material of every organism. In nature, ribosomes use 20 native amino acids to synthesize proteins in most circumstances. However, laboratory efforts to expand the genetic repertoire of living cells and organisms have successfully encoded more than 80 nonnative amino acids in E. coli, yeast, and other eukaryotic systems. The selectivity, fidelity, and site-specificity provided by the technology have enabled unprecedented flexibility in manipulating protein sequences and functions in cells. Various biophysical probes can be chemically conjugated or directly incorporated at specific residues in proteins, and corresponding analytical techniques can then be used to answer diverse biological questions. This review summarizes the methodology of genetic code expansion and its recent progress, and discusses the applications of commonly used analytical methods.