Practical Approach Series – serie

Fermentation is important in the areas of both academic and industrial research. This book is a collection of methods and techniques covering the setting up and use of fermentation units. The emphasis is on the use of small-scale fermenters and the interdisciplinary nature of fermentation itself. As well as being of widespread interest to academic researchers, this technology has many important industrial applications, as evidenced by the affiliations of the authors (Hoechst, ICI, and Beecham Pharmaceuticals).

The most common aim of a biosensor is to produce a signal proportional in strength or frequency to the concentration of chemical or biochemical to which the biological element reacts. This volume describes the design, construction, optimization, and application of the biological component, its attachment to the transducer, and interfacing of the sensing element to appropriate electronic circuitry.

This book is the final part of a trilogy of volumes covering the practical aspects of ligand interactions, biochemistry, and effector mechanisms of signal-transducing receptors. The book focuses on the measurement of receptor binding interactions both at equilibrium and from the kinetic point of view. It includes descriptions of the most important techniques for the performance of ligand binding assays to both membrane-bound and solubilized receptors. In addition to which, a systematic effort has been made to provide insight into the strategy and tactics of receptor binding assays, and the conditions which must be satisfied if artifacts are to be avoided and interpretable data obtained. Since the behaviour of even the simplest receptor systems can be surprisingly complex, an attempt has been made to provide some computational tools which allow complications to be predicted and handled or avoided. Ligand binding assays are very widely used, not least in drug screening and development. This volume will help the practitioner to choose an appropriate ligand, make a suitable receptor preparation, design and perform a well thought out set of assays, and analyse the data obtained correctly.

Advances in the understanding of cellular processes at the molecular level have transformed the biological sciences. Plant pathology is a field which has benefited dramatically from the application of molecular techniques and has become one of the most exciting areas of biology today. Molecular Plant Pathology: A Practical Approach Volume I and its companion Volume II provide the first comprehensive guide to the latest molecular techniques as well as the established approaches to the subject. Detailed protocols are included which address a wide range of investigations from plant pathogen isolation and culture, through physiology and biochemistry, to techniques for localizing genes and their products within the cells of the infected plants. Researchers in plant science, molecular biology, and related areas will find the two volumes of Molecular Plant Pathology to be an invaluable experimental resource, providing a wealth of easy-to-follow protocols supported by expert advice and guidance.

Protein engineering is the rational modification or redesign of proteins using genetic engineering. Thus, it is now possible to modify enzyme specifities, remodel antibodies, and redesign many multi-domain proteins for therapeutic purposes. While the procedures for the introduction of mutations have become routine, predicitng and understanding the effects of these mutations can be complicated. This volume provides a comprehensive guide to the methods used at every stage of the engineering process, from the choice of mutation strategy, through protein stability studies, to critical evaluations of mammalian, yeast, and bacterial host expression systems. Protein Engineering: A Practical Approach is the first practical guide to this fascinating mixture of molecular biology, protein structure analysis, computation, and biochemistry. It combines a thorough theoretical foundation with detailed protocols and will be invaluable to all research workers in the area, from graduate students to senior investigators.

Although most often considered in terms of the generation of action potentials, there are several other electrical, metabolic and ionic correlates of neuronal activity. This book draws together in a single volume, ten different facets of the activity of nerve cells. There are chapters on the monitoring of extracellular unit activity, intracellular voltage and current clamping, patch clamping, video imaging, in vivo voltammetry, intracerebral dialysis, antibody micro- and ion-selective electrodes. The techniques described range from the well-established to the experimental. In each case the chapters include up-to-date information, methodological tips and detailed protocols allowing the reader to follow each method in a stepwise fashion. The wide range of techniques in the book should be of interest to neuroscientists, whether concerned with traditional or novel indices on neuronal function.

Anaerobes are diverse and fascinating organisms. Due to recent advances in equipment and methodology researchers are now able to make increasingly widespread studies of their activities. As a consequence, interest in anaerobes is growing rapidly. This book provides the first ready source of general methods of use to all anaerobic microbiologists, in addition to those which are valuable in the study of specific habitats or groups of organisms. Topics include isolation and culture techniques as well as a range of taxonomic methods, and protocols for biochemical and genetic studies. Several chapters are also devoted specifically to organisms with special culture requirements, such as the methanogens, sulphate-reducing bacteria, and anaerobic eukaryotes (including both protozoa and fungi). This volume will be essential to all those interested in the study of anaerobic micro-organisms, whether in the fields of taxonomy, ecology, physiology or molecular biology.

Acquired chromosome abnormalities are associated with malignancy, exposure to mutagenic agents, and some chromosome instability syndomes. Their study often entails extensive modification of standard cytogenetic techniques as well as a very different approach to analysis. Human Cytogenetics: A Practical Approach Volume II is the first comprehensive manual of techniques for the study of acquired chromosome abnormalities, and as such will be an invaluable source of information for cytogeneticists, haematologists, and oncologists alike.Since the interpretation of chromosome preparations is notoriously difficult in some of these conditions, illustrations, both diagrammatic and photographic, of all chromosome rearrangements discussed are an important feature of this book. In addition to basic techniques, the book takes a futuristic look at new technology which is becoming important in the study of cytogenetics, particularly in the field of cancer research.

This is a concise yet comprehensive practical manual, enabling all those involved in the diagnosis of disease, caused by parasites, to provide both a competent and up-to-date service. There is also sufficient information on the biology of parasites provided to ensure that the book is a valuable resource for educational courses teaching parasitology in both the developed and devloping Worlds.

The topics collected in this volume present classical and molecular techniques currently used in medical virology for the diagnosis of viral infections and for further analysis of virus-specific antibodies and of clinical virus isolates.

Histocompatibility Testing: A Practical Approach describes methods for the investigation of Mhc at the gene and protein levels and their application to the clinical transplanation of human organs and bone marrow. There is currently no other text available covering up-to-date serological and molecular techniques in this area of immunogenetics, despite their importance to fundamental research, clinical practice, and forensic medicine.

Plant cell culture techniques are being increasingly used in basic research and for plant exploitation in industry (genetic engineering, micropropagation). In this book, internationally recognized experts describe the techniques in use in their laboratories for isolating and manipulating cell cultures, and their central importance in plant biotechnology.

Neurochemistry is the study of the cellular and subcellular components of the nervous system and the molecules found within it. The nervous system is complex and neurochemistry a diverse field in which there has been much interest recently, for example in the increased awareness of pathological brain conditions such as Alzheimer's disease, depression, and mania. A wide range of techniques are avialable to the neurochemist and this volume contains the core elements of neurochemistry that will be useful to both first time researchers and established investigators in all fields of neuroscience.

The involvement of carbohydrates in biological processes has greatly fuelled the current interest in this diverse range of molecules. This book presents the up-to-date techniques required to analyse a wide variety of carbohydrates and carbohydrate-containing molecules. This book is intended for all those in the biological/biochemical sciences working on the biological roles and structures of carbohydrates who need detailed protocols for their analysis.

This volume provides detailed laboratory protocols, advice, hints and tips, example data, key literature citations, background information, and troubleshooting comments for the steps used in most molecular biology laboratories. It covers transformation, construction of cDNA and genomic libraries, probe preparation, analysis of gene organisation and expression, in vitro mutagenesis, and DNA sequencing. The information provided is a completely up to date account of each topic by established researchers. These two volumes, the first of four, are a thoroughly revised and up dated version of the original ones published in the Practial Approach series. Together they will offer a complete guide to the major techniques required by modern molecular biology laboratories.

Expression systems are in widespread use for cloning specific genes, for synthesising the encoded proteins for structural and functional analysis, and for large-scale preparation of proteins of commercial interest. This book covers all the main expression systems in current use with background information and advice on the merits of each, step-by-step practical protocols, troubleshooting, and details of the latest applications.

The complexity and cellular heterogeneity of the vertebrate nervous system makes it difficult to study neural processes at the cellular and molecular levels. The establishment of cell culture model systems for almost every region and cell type in the nervous system has therefore been crucial in facilitating such studies and in heralding some key breakthroughs in our understanding of the underlying mechanisms. Neural Cell Culture: A Practical Approach brings together methods for preparing cultures from a diverse range of neural tissues and cells, from both the central and peripheral nervous systems of mammalian and avain species. Important features of this volume are the clearly illustrated dissection guides accompanying each chapter, methods for enriching and purifying distinct cell types, and guidance on optimizing cell viability and growth. Each chapter includes valuable step-by-step protocols, hints, and tips for success, advice, and background information.

Many investigations into the structure and function of cells and tissues require the isolation of a particular membrane or subcellular component (organelle). This book covers all the necessary aspects, from breaking up the cells (homogenization), via a variety of separation techniques (the isolation and fractionation chapters), to characterization of the separated organelles.

The three-dimensional structure of proteins is a key factor in their biological activity. There is an increasing need to be able to predict the structure of a protein once its amino-acid sequence is known; this book presents practical methods of achieving that ambitious aim, using the latest computer modelling algorithms. This work is aimed at postgraduates and postdoctoral researchers in protein science and related fields.

Recombinant DNA techniques have revolutionized the isolation and production of antibodies in recent years, and rapid changes have been made. This new Practical Approach book details protocols which allow the researcher to isolate a new antibody, analyse its properties, format the right antibody molecule or fragment, and produce it in usable quantities.

New and updated, this second edition of Protein Structure presents electrophoretic, chromatographic, and spectrophotometric techniques as well as including new chapters on the relatively sophisticated methods of mass spectrometry and ultracentrifugation. The emphasis is not on the technical aspects of the instruments, but more on the practical aspects of sample preparation and the possibilities of experimental strategy and design. The companion volume to this book, Protein Function: A Practical Approach, details comparable simple techniques for characterizing protein function. Topics in this second volume include: protein molecular weight determination by SDS-PAGE; protein analysis by mass spectrometry; immunological detection of proteins of known sequence; identification of common post-translational modifications; peptide mapping; counting integral numbers of residues by chemical modification; disulfide bonds between cysteine residues; analysis of protein conformation by gel electrophoresis; hydrodynamic properties of proteins; determining the molar absorbance coefficient of a protein; optical spectroscopy to characterize protein conformation and conformational changes; measuring the conformational stability of a protein; imunochemical analysis of protein conformation; stabilization of protein structure by solvents. Protein Structure: A Practical Approach will prove invaluable to all researchers, from PhD students to experienced group leaders.

Protein Expression: A Practical Approach and its companion volume Post-translational Modification: A Practical Approach complete the mini-series of Practical Approach books covering the synthesis and subsequent processing of proteins. Protein Expression: A Practical Approach details the expression of cloned DNA or RNA templates in all the major in vivo and in vitro systems. The in vivo systems covered are cultured mammalian cells, the yeasts Saccharomyces cerevisiae and Pichia pastoris, baculovirus, Xenopus oocytes, and prokaryotic cells. Cell-free systems of both eukarytotes and prokaryotes are described, including the prokaryotic systems that offer copuled transcription- translation. There is also a chapter on monitoring protein expression. The post- translational fate of proteins is covered in Post-Translational Processing: A Practical Approach.

Biomolecules and cells are critical components of biosensors and biomaterials, but in order to function in an artificial environment, they must be immobilized in a manner that does not affect their interaction with target analytes. Biosensors demonstrate that we can harness the incredible functions of living molecules and cells for our own purposes and are therefore at the forefront of technology. Moreover the applications of immobilized biomolecules and cells are expected to expand far beyond biosensor applications and indeed are already used for pharmaceutical production and testing. Biomaterials will become increasing common as they are being developed into toxic filters, artificial organs, and even silicon chips. This book provides a selection of methods for the immobilization of biomolecules and cells on a variety of surface with different geometries and chemistries so that they retain their function and guidelines on which method to use. Also included are the analytical techniques to measure the functionality of immobilized biomolecules. All the protocols have been tried and validated by the authors. Immobilized Biomolecules in Analysis: A Practical Approach is an invaluable guide to all researchers in the fields of biosensors and biomaterials. Research in biosensors is carried out in a wide variety of fields including biochemistry, chemistry, engineering, laboratory medicine, environmental and defence research. The protocols are written so that an extensive prior knowledge of biochemistry is not required to use them.

This new edition of Gel Electrophoresis of Proteins is a completely new text, with eight of the ten chapters written by new authors. It presents the best methods, hints and tips for core procedures such as one- dimensional polyacrylamide gel electrophoresis, isoelectric focusing, two-dimensional gel electrophoresis, preparative gel electrophoresis, and peptide mapping, complete with the latest refinements and updates of the procedures. In addition, it describes major new techniques which have come to the fore since the previous edition. Thus there are chapters on capillary gel electrophoresis, sequence analysis of gel-resolved proteins, fluorophore-labelled saccharide electrophoresis, and analysis of protein:protein interactions by gel electrophoresis. One thing has not changed. The emphasis is still on describing the best methods, in step-by-step detail, with copious advice to ensure that each method works first time in the reader's hands. The first two editions of Gel Electrophoresis of Proteins: A Practical Approach each gained a strong reputation as easy-to-follow laboratory manuals written by experienced researchers for researchers. The methods were presented in a clear accessible format and had been fully tested to ensure success in the lab. This new edition will strengthen the reputation of the book still further. It is a 'must have' for all those who currently use gel electrophoresis or who plan to do so.

Since the first edition of "Liposomes - A Practical Approach" was published in 1990, Liposome technology has become a highly successful and rapidly developing area of preclinical and clinical research. During the 90's, a number of liposome-based drugs reached the market in the U.S. and Europe and several new aspects of liposome research have been discovered. As a consequence there has been an explosion of new procedures and techniques in the field. Reflecting both the significant changes in the field and the new discoveries, this new edition of Liposomes contains a combination of chapters from the 1st edition which have been significantly updated, and several new chapters.Each chapter is an outstanding contribution from leading experts in the field, and the book as a whole encompases all key areas of liposome technology. The first half of the book addresses a number of essential experimental protocols such as the preparation, characterization, and storage of liposomes. Also included are methods for drug encapsulation into liposomes, modification of the liposomal surface to control drug behaviour in biological environments, long-circulating liposomes and the use of cationic liposomes as transfection vectors. The second half of the book discusses specific areas of liposome research and application, such as pH-sensitive liposomes, liposomes for diagnostic imaging, liposomal DNA vaccines, isothermic titration calorimetry of liposomes and vesicular phospholipids gels. This new edition of Liposomes is an essential tool for all experienced liposomologists and the perfect introduction for researchers new to the field.

Drosophila is the scientist's favourite lab organism because it reproduces quickly and has straightforward chromosomes. It has long been the organism of choice in experiments in genetics and is also popular in developmental and behavioural studies. The second edition of this popular Practical Approach book brings the methodology up to date, with contributions from the world's leading Drosophila researchers. It will be valuable to the novice and experienced worker alike.

Like the popular first edition, this new edition of Proteolytic Enzymes emphasizes practical aspects of the handling, characterization, inhibition, and use of proteolytic enzymes giving general advice and specific examples. The text and protocols have been thoroughly updated to take account of the advances made in the last 10 years in both the increased understanding of the role of peptidases in many critical cellular processes e.g. apoptosis and new technological developments e.g. in recombinant protein expression, protein sequencing, and structural studies. The topics covered are: nomenclature and classification; purification; assay methods; determination of mechanism; inhibition and prevention of unwanted proteolytic activity; characterizing natural inhibitors; proteolytic enzymes in peptide mapping and primary structure elucidation by mass spectrometry and Edman sequencing; limited proteolysis as a structural probe; synthetic function. This book will be as invaluable as the first edition in providing ideas and protocols for scientists either studying proteases or using proteases as a research tool.

Proteins are an integral part of molecular and cellular structure and function and are probably the most purified type of biological molecule. In order to elucidate the structure and function of any protein it is first necessary to purify it. Protein purification techniques have evolved over the past ten years with improvements in equipment control, automation, and separation materials, and the introduction of new techniques such as affinity membranes and expanded beds. These developments have reduced the workload involved in protein purification, but there is still a need to consider how unit operations linked together to form a purification strategy, which can be scaled up if necessary. The two Practical Approach books on protein purification have therefore been thoroughly updated and rewritten where necessary. The core of both books is the provision of detailed practical guidelines aimed particularly at laboratory scale purification. Information on scale-up considerations is given where appropriate. The books are not comprehensive but do cover the major laboratory techniques and common sources of protein. Protein Purification Techniques focuses on unit operations and analytical techniques. It starts with an overview of purification strategy and then covers initial extraction and clarification techniques. The rest of the book concentrates on different purification methods with the emphasis being on chromatography. The final chapter considers general scale-up considerations. Protein Purification Applications describes purification strategies from common sources: mammalian cell culture, microbial cell culture, milk, animal tissue, and plant tissue. It also includes chapters on purification of inclusion bodies, fusion proteins, and purification for crystallography. A purification strategy that can produce a highly pure single protein from a crude mixture of proteins, carbohydrates, lipids, and cell debris to is a work of art to be admired. These books (available individually or as a set)are designed to give the laboratory worker the information needed to undertake the challenge of designing such a strategy.

Proteins are an integral part of molecular and cellular structure and function and are probably the most purified type of biological molecule. In order to elucidate the structure and function of any protein it is first necessary to purify it. Protein purification techniques have evolved over the past ten years with improvements in equipment control, automation, and separation materials, and the introduction of new techniques such as affinity membranes and expanded beds. These developments have reduced the workload involved in protein purification, but there is still a need to consider how unit operations linked together to form a purification strategy, which can be scaled up if necessary. The two Practical Approach books on protein purification have therefore been thoroughly updated and rewritten where necessary. The core of both books is the provision of detailed practical guidelines aimed particularly at laboratory scale purification. Information on scale-up considerations is given where appropriate. The books are not comprehensive but do cover the major laboratory techniques and common sources of protein. Protein Purification Techniques focuses on unit operations and analytical techniques. It starts with an overview of purification strategy and then covers initial extraction and clarification techniques. The rest of the book concentrates on different purification methods with the emphasis being on chromatography. The final chapter considers general scale-up considerations. Protein Purification Applications describes purification strategies from common sources: mammalian cell culture, microbial cell culture, milk, animal tissue, and plant tissue. It also includes chapters on purification of inclusion bodies, fusion proteins, and purification for crystallography. A purification strategy that can produce a highly pure single protein from a crude mixture of proteins, carbohydrates, lipids, and cell debris to is a work of art to be admired. These books (available individually or as a set)are designed to give the laboratory worker the information needed to undertake the challenge of designing such a strategy.