PRINCIPLES AND PRACTICE – serie

This invaluable guide to one of the fundamental monitoring techniques in anaesthesia has been updated throughout, and a new chapter has been added on high altitude and flying. Written by a leading recognised expert in the field, this is an authoritative yet clear and practical guide for all on the surgical ward and critical care unit.

Following the succesful publication of "Proteome and Protein Analysis" in 2000, which was based on a former MPSA (Methods in Protein Structure Analysis) conference, Methods in Proteome and Protein Analysis presents the most interesting papers from the 14th MPSA meeting.Major topics include: protein and peptide sample preparation and separation; new reagent for protein sequence analysis; mass spectrometry in protein research; analysis of posttranslational modification; protein-protein interaction using MALDI-MS; manipulation of genome or functional compositon trap; structure-function correlation study using optical biosensors of microcolorimetrical techniques; structural proteomics as NMR or fluorescence polarization study; the classification and prediction of structure or functional sites; in silico analysis of proteins and proteomes; increasing throughput and data quality for proteomics.

The book provides an updated overview of molecular analysis of human tissues, and the impact this analysis has on diagnosis and prognosis of human diseases. Special emphasis is placed on human cancer and the future directions of the field. Methods of handling clinical tissue samples, including the impact of handling on subsequent molecular analysis, are also discussed. In addition, detailed protocols for molecular analysis of DNA, RNA and protein, with special emphasis on molecular analysis of highly complex human tissue samples containing mixtures of cell populations, are provided.

Mammalian and Avian Transgenesis presents a collection of novel methods for the production of a wide range of transgenic animals. The manual focuses largely on mice, but also contains protocols for successful transgenesis in rats, cows, pigs and birds. The manual provides detailed, step-by-step protocols covering all aspects of the production of transgenic animals, including the use of lentiviral vectors in gene transfer, intracytoplasmic sperm injection, nuclear transfer, large insert transgenesis, conditional gene expression systems, the use of reporter genes in transgenesis and transgenesis in large animals and birds. The text is supplemented by superb color photos. While the focus is on newly established techniques, the fundamental methods of transgenesis are also covered for those new to the field. Thus this manual is perfectly suited for those wishing to adopt new technologies in transgenesis.

The adsorption of proteins at interfaces plays a role in many ?elds, such as health, food, environment and analysis. Fundamental aspects are useful when considering applications. We focus here especially on solid-liquid interfaces and present a few fundamental studies regarding adsorption - netics and conformational changes, and examples of applications to sensors and membranes. The ?rst part is dedicated to fundamental studies performed using - tical waveguide lightmode spectroscopy, as an example of a technique that has the advantage of not requiring labelled proteins, but is limited to s- ci?c supports. Conversely, the radiolabelling of proteins, which has the disadvantage of any labelling process, allows application to any kind of s- faces. As proteins bear both positive and negative charges, we can expect thein?uenceofanelectric?eldnormaltothe interfaceonthe pack- ing order at interfaces. The re?ning of data treatment may also lead to the determination of useful structural parameters. The balance between protein-surface and protein-protein interactions is a key point for the - scription of the structure at high coverage of the surface.Electrokinetic methods, like measurement of the streaming potential, may be helpful in the electrical characterisation of the interfacial layer facing the solution. The second part includes different bench techniques that were dev- oped to improve the sensitivity of the characterisation of the orientation and structure of the proteins at interfaces: dual polarisation interferometry and total internal re?ection ellipsometry are such recent examples.

This volume describes high-throughput approaches to a series of robust, established methodologies in molecular genetic studies of population samples. Such developments have been essential not only to linkage and association studies of single-gene and complex traits in humans, animals and plants, but also to the characterisation of clone banks, for example in mapping of genomes. Chapters have been written by developers or highly experienced end-users concerned with a diverse array of biological applications. The book should appeal to any researcher for whom costs and throughput in their genetics laboratory have become an issue.

This volume describes high-throughput approaches to a series of robust, established methodologies in molecular genetic studies of population samples. Such developments have been essential not only to linkage and association studies of single-gene and complex traits in humans, animals and plants, but also to the characterisation of clone banks, for example in mapping of genomes. Chapters have been written by developers or highly experienced end-users concerned with a diverse array of biological applications. The book should appeal to any researcher for whom costs and throughput in their genetics laboratory have become an issue.

This complete and practical manual on expression measurement using DNA arrays covers the existing methods (from nylon macroarrays to oligonucleotide chips) and includes detailed protocols. It has been written by practising scientists who have experienced the difficulties involved in actually using microarrays, and provides helpful advice and hints on setting up these powerful but sometimes tricky methods. Software, data mining procedures and probable future developments, which should be useful to any practising scientist interested in expression measurement, are also covered in this book. It also provides detailed protocols as well as many helpful hints to achieve experimental success and to avoid pitfalls.

Two-dimensional electrophoresis is the central methodology in proteome research, and the state of the art is described in detail in this text, together with extensive coverage of the detection methods available. Sufficient detail is given to allow the readers to apply these technologies to their own particular requirements.

Two-dimensional electrophoresis is the central methodology in proteome research, and the state of the art is described in detail in this text, together with extensive coverage of the detection methods available. Sufficient detail is given to allow the readers to apply these technologies to their own particular requirements.

A better "casting" could not be conceived. The authors of this book are gold­ smiths on the subject. I have followed their work since their "entry" into cyto­ genetics and I have a high esteem for them. I consider it an honour to be asked to write the preface of their opus. Paul Popescu, Directeur de Recherche at INRA, has also played a promi­ nent part in the development of animal cytogenetics, especially in domestic animals. He is able to tell you the cost of a translocation in a pig breeding farm or a cow population: a fortune! P. Popescu has played a great part in gene mapping of these species using "in situ DNA hybridisation". His contributions are recognised world-wide. His laboratory receives many visitors every year and it serves as a reference for domestic animal cytogenetics. Helene Hayes, Charge de Recherche at INRA, has collaborated with P. POPESCU in the elaboration of the "at hand" techniques and in many other discoveries which are listed in her bibliography. She showed the fascinating correspondence between bovine and human chromosomes and the com­ pared gene maps of domestic bovidae.

Recent advances in large-scale DNA sequencing technology have made it possible to sequence the entire genome of an organism. Attention is now turning to the analysis of the product of the genome, the proteome, which is the set of proteins being expressed by a cell. Two-dimensional gel electrophoresis can be used to create cellular protein maps which give a quantitative and qualitative picture of the proteome. Mass spectrometry is the method of choice for the rapid large-scale idenfification of these proteomes and their modifications. An understanding of these methods is critical for scientists in the "Post-Genome" era.

Recent advances in large-scale DNA sequencing technology have made it possible to sequence the entire genome of an organism. Attention is now turning to the analysis of the product of the genome, the proteome, which is the set of proteins being expressed by a cell. Two-dimensional gel electrophoresis can be used to create cellular protein maps which give a quantitative and qualitative picture of the proteome. Mass spectrometry is the method of choice for the rapid large-scale idenfification of these proteomes and their modifications. An understanding of these methods is critical for scientists in the "Post-Genome" era.

The completion of the Human Genome Project and the rapid progress in cell bi- ogy and biochemical engineering, are major forces driving the steady increase of approved biotech products, especially biopharmaceuticals, in the market. Today mammalian cell products (“products from cells”), primarily monoclonals, cytokines, recombinant glycoproteins, and, increasingly, vaccines, dominate the biopharmaceutical industry. Moreover, a small number of products consisting of in vitro cultivated cells (“cells as product”) for regenerative medicine have also been introduced in the market. Their efficient production requires comprehensive knowledge of biological as well as biochemical mammalian cell culture fundamentals (e.g., cell characteristics and metabolism, cell line establishment, culture medium optimization) and related engineering principles (e.g., bioreactor design, process scale-up and optimization). In addition, new developments focusing on cell line development, animal-free c- ture media, disposables and the implications of changing processes (multi-purpo- facilities) have to be taken into account. While a number of excellent books treating the basic methods and applications of mammalian cell culture technology have been published, only little attention has been afforded to their engineering aspects. The aim of this book is to make a contribution to closing this gap; it particularly focuses on the interactions between biological and biochemical and engineering principles in processes derived from cell cultures. It is not intended to give a c- prehensive overview of the literature. This has been done extensively elsewhere.

Proteomics is a multifaceted, interdisciplinary field which studies the complexity and dynamics of proteins in biological systems. It combines powerful separation and analytical technology with advanced informatics to understand the function of proteins in the cell and in the body. This book provides a clear conceptual description of each facet of proteomics, describes recent advances in technology and thinking in each area, and provides details of how these have been applied to a variety of biological problems. It is written by expert practitioners in the field, from industry, research institutions, and the clinic. It provides junior and experienced researchers with an invaluable proteomic reference, and gives fascinating glimpses of the future of this dynamic field.

“Imaging Cellular and Molecular Biological Function” provides a unique selection of essays by leading experts, aiming at scientist and student alike who are interested in all aspects of modern imaging, from its application and up-scaling to its development. The chapter content ranges from the basic through to complex overview of method and protocols, and there is also practical and detailed “how-to” content on important, but rarely addressed topics. This first edition features all-colour-plate chapters. The philosophy of this volume is to provide student, researcher, PI, professional or provost the means to enter this applications field with confidence, and to construct the means to answer their own specific questions.

The completion of the Human Genome Project and the rapid progress in cell bi- ogy and biochemical engineering, are major forces driving the steady increase of approved biotech products, especially biopharmaceuticals, in the market. Today mammalian cell products (“products from cells”), primarily monoclonals, cytokines, recombinant glycoproteins, and, increasingly, vaccines, dominate the biopharmaceutical industry. Moreover, a small number of products consisting of in vitro cultivated cells (“cells as product”) for regenerative medicine have also been introduced in the market. Their efficient production requires comprehensive knowledge of biological as well as biochemical mammalian cell culture fundamentals (e.g., cell characteristics and metabolism, cell line establishment, culture medium optimization) and related engineering principles (e.g., bioreactor design, process scale-up and optimization). In addition, new developments focusing on cell line development, animal-free c- ture media, disposables and the implications of changing processes (multi-purpo- facilities) have to be taken into account. While a number of excellent books treating the basic methods and applications of mammalian cell culture technology have been published, only little attention has been afforded to their engineering aspects. The aim of this book is to make a contribution to closing this gap; it particularly focuses on the interactions between biological and biochemical and engineering principles in processes derived from cell cultures. It is not intended to give a c- prehensive overview of the literature. This has been done extensively elsewhere.

Following the succesful publication of "Proteome and Protein Analysis" in 2000, which was based on a former MPSA (Methods in Protein Structure Analysis) conference, Methods in Proteome and Protein Analysis presents the most interesting papers from the 14th MPSA meeting.Major topics include: protein and peptide sample preparation and separation; new reagent for protein sequence analysis; mass spectrometry in protein research; analysis of posttranslational modification; protein-protein interaction using MALDI-MS; manipulation of genome or functional compositon trap; structure-function correlation study using optical biosensors of microcolorimetrical techniques; structural proteomics as NMR or fluorescence polarization study; the classification and prediction of structure or functional sites; in silico analysis of proteins and proteomes; increasing throughput and data quality for proteomics.

The book provides an updated overview of molecular analysis of human tissues, and the impact this analysis has on diagnosis and prognosis of human diseases. Special emphasis is placed on human cancer and the future directions of the field. Methods of handling clinical tissue samples, including the impact of handling on subsequent molecular analysis, are also discussed. In addition, detailed protocols for molecular analysis of DNA, RNA and protein, with special emphasis on molecular analysis of highly complex human tissue samples containing mixtures of cell populations, are provided.

Mammalian and Avian Transgenesis presents a collection of novel methods for the production of a wide range of transgenic animals. The manual focuses largely on mice, but also contains protocols for successful transgenesis in rats, cows, pigs and birds. The manual provides detailed, step-by-step protocols covering all aspects of the production of transgenic animals, including the use of lentiviral vectors in gene transfer, intracytoplasmic sperm injection, nuclear transfer, large insert transgenesis, conditional gene expression systems, the use of reporter genes in transgenesis and transgenesis in large animals and birds. The text is supplemented by superb color photos. While the focus is on newly established techniques, the fundamental methods of transgenesis are also covered for those new to the field. Thus this manual is perfectly suited for those wishing to adopt new technologies in transgenesis.

The adsorption of proteins at interfaces plays a role in many ?elds, such as health, food, environment and analysis. Fundamental aspects are useful when considering applications. We focus here especially on solid-liquid interfaces and present a few fundamental studies regarding adsorption - netics and conformational changes, and examples of applications to sensors and membranes. The ?rst part is dedicated to fundamental studies performed using - tical waveguide lightmode spectroscopy, as an example of a technique that has the advantage of not requiring labelled proteins, but is limited to s- ci?c supports. Conversely, the radiolabelling of proteins, which has the disadvantage of any labelling process, allows application to any kind of s- faces. As proteins bear both positive and negative charges, we can expect thein?uenceofanelectric?eldnormaltothe interfaceonthe pack- ing order at interfaces. The re?ning of data treatment may also lead to the determination of useful structural parameters. The balance between protein-surface and protein-protein interactions is a key point for the - scription of the structure at high coverage of the surface.Electrokinetic methods, like measurement of the streaming potential, may be helpful in the electrical characterisation of the interfacial layer facing the solution. The second part includes different bench techniques that were dev- oped to improve the sensitivity of the characterisation of the orientation and structure of the proteins at interfaces: dual polarisation interferometry and total internal re?ection ellipsometry are such recent examples.

“Imaging Cellular and Molecular Biological Function” provides a unique selection of essays by leading experts, aiming at scientist and student alike who are interested in all aspects of modern imaging, from its application and up-scaling to its development. The chapter content ranges from the basic through to complex overview of method and protocols, and there is also practical and detailed “how-to” content on important, but rarely addressed topics. This first edition features all-colour-plate chapters. The philosophy of this volume is to provide student, researcher, PI, professional or provost the means to enter this applications field with confidence, and to construct the means to answer their own specific questions.

This book provides a state-of-the-art approach to the molecular basis of hematologic diseases and its translation into improved diagnostics and novel therapeutic strategies. Several representative hemato-oncologic malignancies are analyzed in detail: acute lymphoblastic leukemia, acute myeloid leukemia, B-cell Non-Hodgkin lymphomas, multiple myeloma, chronic lymphocytic leukemia, chronic myeloid leukemia, myelodysplastic syndromes, and myeloproliferative neoplasms. Experts in the field describe the molecular methods applied for modern diagnostics and therapies, such as hematopoietic stem cell transplantation, donor recipient matching, banking of biological material, analyses of post-transplant chimerism, and minimal residual disease monitoring. The volume concludes with an extensive section comprising thorough step-by-step protocols of molecular techniques in hematology, all of them validated in the authors’ own laboratories.

Proteomics is a multifaceted, interdisciplinary field which studies the complexity and dynamics of proteins in biological systems. It combines powerful separation and analytical technology with advanced informatics to understand the function of proteins in the cell and in the body. This book provides a clear conceptual description of each facet of proteomics, describes recent advances in technology and thinking in each area, and provides details of how these have been applied to a variety of biological problems. It is written by expert practitioners in the field, from industry, research institutions, and the clinic. It provides junior and experienced researchers with an invaluable proteomic reference, and gives fascinating glimpses of the future of this dynamic field.

This book provides a state-of-the-art approach to the molecular basis of hematologic diseases and its translation into improved diagnostics and novel therapeutic strategies. Several representative hemato-oncologic malignancies are analyzed in detail: acute lymphoblastic leukemia, acute myeloid leukemia, B-cell Non-Hodgkin lymphomas, multiple myeloma, chronic lymphocytic leukemia, chronic myeloid leukemia, myelodysplastic syndromes, and myeloproliferative neoplasms. Experts in the field describe the molecular methods applied for modern diagnostics and therapies, such as hematopoietic stem cell transplantation, donor recipient matching, banking of biological material, analyses of post-transplant chimerism, and minimal residual disease monitoring. The volume concludes with an extensive section comprising thorough step-by-step protocols of molecular techniques in hematology, all of them validated in the authors’ own laboratories.

nomenon [26]. Indeed, Krieg et al. [21] showed that the elimination of the CpG in a particular ODN invariably abolished immune stimulation, but changes in the ODN sequences that did not affect the CpG or the flanking bases did not alter the immuno- stimulatory (IS) effect. Furthermore, they extended the initial observations of the IS effects to non-palindromic CpG-enriched ODN [21]. Subsequent studies showed that CpG-enriched ODN also induce the secretion of IL-6 and IL-12 [19] and IFN-a [6, 27]. By adding or deleting various IS sequences (ISS)-ODN to or from different pDNAs, it was demonstrated that the ISS have a pivotal role in the induction of the subsequent immune response to the gene product in gene-vaccinated animals. The enhanced Thl immune response induced by gene vaccination is the consequence of the activation of the innate immune response by the ISS in the pDNA backbone [30, 31], rather than the low dose of intracellularly produced antigen. The cell activation products induced by the ISS, i. e. , IFN-a [3], IFN-~ [43], IL-12 [37], and IL-18 [25], are established inducers of IFN-y synthesis and promote the differentiation of naive T helper cells to Thl lym- phocytes.Thus, the ISS activate the precise innate cytokine network required to pro- mote Thl differentiation (see Fig. 1). In a recent study it was demonstrated that this ap- proach is also applicable to a protein antigen.

A better "casting" could not be conceived. The authors of this book are gold­ smiths on the subject. I have followed their work since their "entry" into cyto­ genetics and I have a high esteem for them. I consider it an honour to be asked to write the preface of their opus. Paul Popescu, Directeur de Recherche at INRA, has also played a promi­ nent part in the development of animal cytogenetics, especially in domestic animals. He is able to tell you the cost of a translocation in a pig breeding farm or a cow population: a fortune! P. Popescu has played a great part in gene mapping of these species using "in situ DNA hybridisation". His contributions are recognised world-wide. His laboratory receives many visitors every year and it serves as a reference for domestic animal cytogenetics. Helene Hayes, Charge de Recherche at INRA, has collaborated with P. POPESCU in the elaboration of the "at hand" techniques and in many other discoveries which are listed in her bibliography. She showed the fascinating correspondence between bovine and human chromosomes and the com­ pared gene maps of domestic bovidae.

In spite of tremendous scientific progress over the past years, cell biologists do not yet understand the fundamental processes that determine the life cy­ cle of a cell. Such are: cell movement and cell spreading, cell division, cell communication, cell signaling, cell regeneration and cell death. Biochemistry has enabled us to recognize and to isolate an overwhelming number of new proteins. In vitro assays and the reinjection of proteins into cells and tissues have provided insights into molecular functions and cellular mechanisms. The renaissance of the genetic approach by applying restriction enzymes and vectors, PCR and antisense technology has enabled us to overexpress certain cellular products, to make altered constructs of cell components or to create "knock-out" mutants that entirely lack the factor of interest. Amazingly en­ ough, all these molecular toys have led to a stream of information but not, in a comparable degree, to a better understanding. Has the puzzle become too complex to get solved; or are the windows too small that we are looking through? As an attempt to answer both questions, the aim of the present mono­ graph Modern Optics, Electronics and High Precision Techniques in Cell Biol­ ogy is first to provide cell and molecular biologists with a whole new scope of easily applicable techniques including brand-new optical, biophysical, physicochemical and biosensoric devices. Secondly, these newly developed techniques allow us to look at cells and biological systems as a whole.