bluuebunny/crossref_metadata_2025_split huggingface dataset:
Polymerase Chain Reaction and Other Amplification Systems
Authors: Christopoulos Theodore K. 
Published: September 2006

Abstract
The discovery of restriction endonucleases is perhaps the most significant technical milestone in the development of recombinant DNA technology. For, together with ligation and transformation procedures, they enabled the insertion of genes of any organism in suitable vectors (e.g. plasmids), introduction of the vector in bacteria and subsequent replication of the DNA of interest during cell division (cloning). Other important technical milestones include the development of methods for DNA sequencing and the chemical synthesis of oligodeoxynucleotides.
The polymerase chain reaction (PCR) is another technique which had a major impact in every area of research that involves nucleic acid analysis. PCR is a method of exponentially increasing the concentration of a specific nucleic acid sequence in a reaction mixture. A great advantage of PCR is that the target nucleic acid does not need to be particularly pure. It can be a minute part of an extremely complex mixture of biological material. Crude cell lysates provide a very good starting material. The ability to propagate, in a few hours, minute amounts of DNA that are too small for standard amplification (i.e. cloning) techniques gives PCR an extraordinary power and sensitivity.
The advent of PCR stimulated the research activity in the development of alternative exponential amplification systems such as the ligase chain reaction (LCR), the self‐sustained sequence replication (3SR), the nucleic acid sequence‐based amplification (NASBA) and the strand displacement amplification system (SDA). An essential feature of target amplification techniques is that the amplification is an iterative process consisting of successive cycles. The products synthesized in one cycle act as templates for the next cycle.
Amplification techniques can be used to amplify trace amounts of nucleic acid sequences in blood, cells, water, food and other clinical and environmental samples. Fixed tissue specimens, buccal cells from mouth washes, human hair, dried blood at the scene of a crime, single lymphoid or sperm cells are examples of starting material that may be used for nucleic acid analysis.
Radiation Hybrid Mapping
Authors: Newell William R. 
Published: September 2006

AbstractRadiation hybrid (RH) mapping is a convenient and widely used method for rapidly locating new DNA sequences on genomes in relation to other DNA markers. The aim of the technique is to determine the correct order and relative distances between a set of markers, or to locate a new marker to its most likely location on a genome. We review the history of the technique, its current status, and the main analytical techniques used to generate maps from raw data. The physical processes and assumptions underlying the technique are examined. These processes are used to derive models from which the required information, location of markers with respect to each other on a linear map of DNA, is obtained by a variety of computational means. The RH technique has proved indispensable in mapping the human genome, since, unlike genetic mapping, the conditions under which panels are made is completely under experimental control, so mapping at any degree of resolution is possible. The other major advantage of RH mapping over genetic mapping is that markers need not be polymorphic; it is presence or absence of a marker that is detected, not type. It has therefore been used to map the large numbers of expressed sequence tags that have been generated as part of the human genome project (HGP). This has given rise to the first human gene map and subsequent updates, so that currently 40 000 genes have been assigned locations on the human genome.
Structural Analysis of Ribozymes
Authors: Ananvoranich Sirinart , Perreault Jean‐Pierre 
Published: September 2006

Abstract
We use delta ribozymes as a model in illustrating the various steps involved in the structural analysis of ribozymes. Delta ribozymes were originally identified as a self‐cleaving motif located on the single‐stranded RNA genome of hepatitis delta virus (HDV). The methodologies are presented in the order of their requirement for determination of the structure/function relationship of a ribozyme, beginning with the preparation of the RNA molecules required for the studies. Primary structural information is needed in order to identify potential ribozymes, then secondary and tertiary structural information is required for ribozyme characterization. The manuscript was written to guide an investigator from the initial observation of RNA catalytic activity (a ribozyme discovery) to the deduction of a structural model of the ribozyme.
Centrifugation in Particle Size Analysis
Authors: Mächtle Walter 
Published: September 2006

Abstract
Millions of tons per year of submicron particles are produced in industrial processes around the world. They are also referred to as nanoparticles, colloids, latices, polymer dispersions, mini‐emulsions or micro‐emulsions. These solid or liquid particles, which are mostly dispersed in an aqueous medium, have a diameter (D) in the 10 < D < 3000 nm range. They are used as dyes and pigments, in paints, adhesives and coatings applied to paper, as impact modifiers in plastics, as catalysts, in cosmetic and photographic emulsions, metal sols and electrodeposition coatings applied to automobiles, as excipients for pharmaceuticals, etc.
Most of the valuable practical properties of these nanoparticle dispersions/emulsions result from their extremely small size and their high specific surface area. This implies that it is essential to be able to control and measure the particle size in order to produce nanoparticle systems of consistent quality. The particle size distribution is just as important as the average particle diameter. The practical consequences of changing from a very narrow to a very broad particle size distribution with the same average diameter can be quite dramatic in many applications. For example, the viscosity of a high‐solid polymer latex will decrease, the impact strength of plastic material will increase, surfaces coated with emulsion paints will become more level, etc.

Many methods, such as light scattering, electron microscopy, field flow fractionation (FFF), capillary hydrodynamic chromatography and ultracentrifugation, are used to measure the average particle diameters and overall particle size distribution of nanoparticles. Analytical ultracentrifugation (AUC) is the most versatile method with the broadest diameter range (1–5000 nm) and the highest resolution. The nanoparticles are forced to move through a predominantly aqueous medium inside an AUC measuring cell under a centrifugal force of 500–500 000 × g (rotor speed 100–60 000 rpm). Sedimentation causes fractionation to take place, because the larger particles move faster. The sedimentation velocities s
i
of the different fractions i are measured with refractive index, ultraviolet (UV)‐absorption or light scattering/turbidity detectors. The diameters D
i
of the different fractions can be calculated from the difference in s
i
, i.e. the complete particle size distribution, according to Stokes' law. The resolution of the AUC method is so high because, according to Stokes, s
i
is proportional to the square of D
i
. The AUC particle size distribution method can give spurious results if D is too high, i.e. if the particles form a sediment too quickly or if they display a broad chemical heterogeneity (i.e. no uniform particle density).
Optical Particle Counting
Authors: Lieberman Alvin 
Published: September 2006

Abstract
This discussion of optical particle counting (OPC) points out application areas where the procedure is used to collect data that would not be available otherwise. These areas include determination of particle size distributions from the size data of individual particles, identification of particles that do not fit within a specified size distribution function, and characterizing the particles present in a process of product fluid. The designs, performance, and operation of OPCs for both gasborne and liquidborne particles are discussed. Capabilities and limitations in terms of particle size ranges, counting accuracy, concentration, and so on are pointed out. The primary limitation is related to the fact that OPCs respond not only to the size of the particles, but also to the optical nature of the particles and of the fluid. Data are also affected by the counter design. All respond to a physical and/or chemical property of the particles, as well as to their physical size and morphology. The limitations of OPCs in relation to those factors are considered.
Operational requirements for OPC are pointed out. These include good sample acquisition and handling procedures, consideration of the effects of differences between particle counter designs on reported data, requirements for careful calibration before use and the effects of differences between calibration particles and sampled particles. A brief summary of present national and international standard documents for calibration and operation of these instruments is included. Performance capabilities of current OPCs are summarized and possible future developments are discussed. Performance is considered in comparison with that of other particle measuring procedures.
Photon Correlation Spectroscopy in Particle Sizing
Authors: Tscharnuter Walther 
Published: September 2006

Abstract
Photon correlation spectroscopy (PCS) has become a powerful light‐scattering technique for studying the properties of suspensions and solutions of colloids, macromolecules and polymers, that is absolute, non‐invasive and non‐destructive. This text explains the principles of the technique and describes the required instrumentation. It also discusses new developments in instrumentation, which may substantially impact on future applications of this technique. Proper sample preparation is of major importance for the accuracy and precision of the mean size and size distribution results and thus one section is dedicated to sample handling.
Sedimentation in Particle Size Analysis
Authors: Bernhardt Claus 
Published: September 2006

AbstractParticle size analysis by sedimentation is one of the most important techniques of analysis. The principle is based on the fact that it is possible to draw conclusions about the geometrical dimensions (size) of the particles from the speed of their movement in a fluid. In the past many different apparatuses were developed. Nowadays, methods that allow quick, automatic measurement are most popular.This article consists of two general sections valid for all methods: the first concerns the undisturbed and disturbed movement of particles in a fluid at rest, and the second concerns the dispersion of solids in liquids.The methods described are divided in gravitational methods and methods used in a centrifugal field, respectively. Finally, the appendix gives a list of liquids and dispersing agents valid for measurement.
Surface Area and Pore Size Distributions
Authors: Takei Takashi 
Published: September 2006

AbstractThe determination methods of the surface area and pore size distribution (PSD) of a solid have been extensively described. The surface area and PSD are important as the basic properties for a powder and porous material. Nowadays the surface area and PSD are easily obtained by an automatic measuring instrument. However, there is no method which covers the complete range of a specific surface area and a pore size. All of the methods have their limits of measurement. In many cases the definitions of the surface area and pore size in each method differ from one another. The features and principles of the methods have been discussed mainly in order to select an appropriate method and to analyze data exactly.
Capillary Electrophoresis of Proteins and Glycoproteins
Authors: Trout Amy , Muza Michelle M. , Landers James P. 
Published: September 2006

AbstractAs a result of their complex nature, the separation of proteins and glycoproteins by capillary electrophoresis (CE) is a challenge. Fortunately, a number of approaches allow for effective analysis of these biomolecules. Of particular concern is the proclivity of proteins and their glycoconjugates to adsorb to the capillary wall. Various covalent and dynamic capillary coatings are described which minimize this effect. We also address general concerns including electrode polarity, capillary conditioning and method development that are important for achieving optimal separation. Finally, each separation mode, including capillary zone electrophoresis (CZE), capillary gel electrophoresis (CGE), and capillary isoelectric focusing (CIEF), is discussed with particular attention to its niche in the realm of protein and glycoprotein analysis.
Fluorescence Spectroscopy in Peptide and Protein Analysis
Authors: Ladokhin Alexey S. 
Published: September 2006

AbstractFluorescence spectroscopy and its multiple applications to the life sciences have undergone rapid development. This is due to numerous technical advances in both instrumentation and methods of data analysis as well as to a vast proliferation of basic techniques. Applications of fluorescence spectroscopy to protein and peptide analysis are governed by three principal factors: the dynamic nature of the signal, its localized nature, and its redundancy. Although these features can complicate interpretation of the experimental result, they also can be exploited to obtain unique structural and dynamic information. The availability and simplicity of basic data acquisition and analysis are important practical features behind the popularity of fluorescence as compared to other spectroscopic techniques. Yet this simplicity does not appear to compromise its advantages. This article is intended first to provide an overview of the fluorescence phenomenon in proteins, and second to illustrate applications of fluorescence spectroscopy in advanced (but not necessarily high‐tech) studies. Three areas of protein studies, namely protein–ligand interactions, protein folding and studies of membrane proteins, have been chosen to demonstrate key advantages of fluorescence spectroscopy: it is sensitive, versatile, and it lends itself readily to fast data acquisition.
Protein Purification: Theoretical and Methodological Considerations
Authors: Takács Béla János 
Published: September 2006

AbstractWith the rapid advances in recombinant DNA technologies, it became possible to express large amounts of virtually any protein in different host cells, including bacteria, yeast, insect, and mammalian cells. Efficient purification methods are becoming more and more important as the need for protein drugs continues to expand. In this article we cover some basic procedures for the purification of soluble recombinant proteins expressed in Escherichia coli and methods to obtain soluble and active protein from insoluble and inactive inclusion bodies. Finally, methods for the purification of native intrinsic membrane proteins from human lymphoblastoid cell lines, using immobilized monoclonal antibodies (mAbs), are presented.As a first step, the protein of interest has to be released into an aqueous extract. Following extraction, residual debris and intact cells are removed by centrifugation. Nucleic acids are either hydrolyzed or precipitated to reduce the viscosity of the extract. A crude enrichment for the protein of interest is made based on its differential solubility in ammonium sulfate. This is then followed by high‐resolution chromatographic methods. The type of chromatographic matrices and the order of their use are evaluated in pilot experiments for column selection. By combining the specificity and discriminatory powers of various chromatographic techniques, it is possible to isolate rapidly a single protein species from a heterogeneous mixture.For the purification of native intrinsic membrane proteins, the use of immunoaffinity methods is emphasized, since specific mAbs may be obtained even against low‐copy‐number membrane proteins by reducing a multispecific antibody response to a series of monospecific responses by cloning. The use of immobilized, highly specific mAbs is demonstrated in the purification of human leukocyte antigen D‐related (HLA‐DR) antigens.Rapid advances in complete genome sequences provide us with a continuously expanding number of new potential therapeutic targets. These proteins will have to be purified and characterized. The existence of a functional protein blueprint will be invaluable not only for the diagnosis but possibly also for the treatment of many diseases.
Proteolytic Mapping
Authors: Nadler Tim 
Published: September 2006

Abstract
Proteolytic mapping is a technique used to verify the primary structure of a protein. It involves the cleavage of the protein at specific amino acids to generate a unique set of peptides which is then separated and analyzed. The chromatogram generated during the separation is known as the proteolytic map of the protein, which serves as a unique “fingerprint” for that protein. A proteolytic map can be used to verify the identity of a protein and to detect mutations in a protein. This technique is faster than two‐dimensional (2D) nuclear magnetic resonance (NMR), X‐ray crystallography, or complete amino acid sequencing for verifying the primary structure of a protein. However, it does not allow determination of the sequence of a protein de novo as can be accomplished with sequencing, nor does it yield the higher order structural information that may be obtained with 2D NMR or X‐ray crystallography.
The process of proteolytic mapping involves sample preparation, enzyme digestion, and map development. Each of these steps should be optimized for each new protein to obtain a good map; alternatively, generic protocols can be used to produce satisfactory results for some proteins. A typical protocol requires about a day to complete owing to the time required to digest a protein with an enzyme. However, more recent automated techniques are capable of completing the process in a few hours because they employ a highly concentrated immobilized enzyme which speeds digestion. Proteolytic maps are used in both quality control (QC) and research environments, with the most recent application in the field of proteomics.
Reversed‐Phase High‐Performance Liquid Chromatography in Peptide and Protein Analysis
Authors: Aguilar Marie‐Isabel 
Published: September 2006

Abstract
Reversed‐phase high‐performance liquid chromatography (RPHPLC) involves the separation of molecules on the basis of hydrophobicity. The separation depends on the hydrophobic binding of the solute molecule from the mobile phase to the immobilized hydrophobic ligands attached to the stationary phase, i.e. the sorbent. The solute mixture is initially applied to the sorbent in the presence of aqueous buffers, and the solutes are eluted by the addition of organic solvent to the mobile phase. Elution can proceed either under isocratic conditions, where the concentration of organic solvent is constant, or by gradient elution, whereby the amount of organic solvent is increased over a period of time. The solutes are therefore eluted in order of increasing molecular hydrophobicity. RPHPLC is a very powerful technique for the analysis of peptides and proteins because of a number of factors which include (1) the excellent resolution that can be achieved under a wide range of chromatographic conditions for very closely related molecules in addition to structurally distinct molecules, (2) the experimental ease with which chromatographic selectivity can be manipulated through changes in mobile phase characteristics, (3) the generally high recoveries and hence high productivity and (4) the excellent reproducibility of repetitive separations carried out over a long period of time, which is due partly to the stability of the sorbent materials under a wide range of mobile phase conditions. However, RPHPLC can cause the irreversible denaturation of protein samples, thereby reducing the potential recovery of material in a biologically active form.
X
‐Ray Crystallography of Biological Macromolecules
Authors: Messerschmidt Albrecht , Huber Robert 
Published: September 2006

Abstract
X‐ray crystallography of biological macromolecules comprises the determination of the three‐dimensional structures of proteins, nucleic acids and other biological macromolecules at atomic resolution by diffraction of X‐rays on crystals of such macromolecules. The atomic structure is obtained from the electron density distribution of the macromolecular crystal, which is the Fourier transform of the waves diffracted by the crystal. The amplitudes of the diffracted waves are determined directly from the diffraction experiment. The diffraction phases are revealed (i) from additional diffraction data of isomorphous heavy atom derivatives [multiple isomorphous replacement (MIR) technique], (ii) from multiple anomalous diffraction (MAD), if suitable anomalous scatterers are in the crystal and using tunable synchrotron radiation, and (iii) by Patterson search techniques (molecular replacement), if structural information on the biological macromolecule under investigation is available. X‐ray crystallography of biological macromolecules is the unique method for the elucidation of the spatial structures at atomic resolution of complex biomolecules with molecular masses greater than 30 kDa. The structures represent a time average and spatial average of molecules packed in a crystal. The preparation of suitable crystals may be a limiting factor.
Biological Matrices: Pesticides Content Sampling, Sample Preparation and Preservation
Authors: Kawasaki Seiji , Tadano Jutaro 
Published: September 2006

AbstractEnvironmental pollution is a world‐wide problem. Consequently, the demand for pesticide analysis in biological samples is increasing. In most cases, the availability of the test result depends on the suitability of sampling, sample preservation, and sample preparation which must satisfy the need. However, there is no reference method. This paper summarizes sampling and sample preparation and preservation methods for the case when the compound to be detected is not known.
Gas Chromatography/Mass Spectrometry Methods in Pesticide Analysis
Authors: Benfenati Emilio , Natangelo Marco , Tavazzi Simona 
Published: September 2006

Abstract

Gas chromatography/mass spectrometry (GC/MS) is irreplaceable in pesticide analysis. Nowadays international bodies consider mass spectrometry (MS) the method of choice for pesticide analysis and quantitative methods using GC/MS are more and more numerous.
1,2
The key characteristics of GC/MS in pesticide analysis are its selectivity and sensitivity, but other important features will be considered in detail in the specific sections on identification and quantitative analysis.

A great advantage of GC/MS in identification is that it gives a mass spectrum that can be easily and quickly compared on‐line with a library of more than 200 000 mass spectra. Specific pesticide libraries are available with hundreds of compounds. Mass spectra can be obtained with just a few picograms of compound injected and gas chromatography (GC) can separate analytes. This is an unbeatable feature of GC/MS.
In quantitative analysis, sensitivity is often excellent and selectivity is better than with other GC detectors. Multiresidue analyses can easily be done. The use of internal standards labeled with stable isotopes allows an accuracy which is not achieved with common methods. These characteristics facilitate sample processing because cleanup stages are not necessary in many cases.
Major limitations in the use of GC/MS depend on GC limits. Thus, very polar and thermally labile pesticides, not suitable for GC analysis, cannot be analyzed by GC/MS. Other limitations are the cost of the apparatus and the relative complexity of the method, compared with the simpler GC detectors. However, these limitations are gradually becoming less important as GC/MS instruments become simpler and cheaper.