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This book is a collection of critical reviews of the use of high-performance liquid chromatography in a very specialized area of research. It describes in detail modern methodology to separate nucleic acids, enzymes and a wide variety of biologically active proteins such as renin.

Intrinsically Disordered Proteins, Volume 611, the latest release in the Methods in Enzymology series, highlights new advances in the field, with this new volume presenting interesting chapters on topics of interest, including the Characterization of Structure-Function relationships in the intrinsically disordered protein complexin, Distances, distance distributions, and ensembles of IDPs from single-molecule FRET, Biophysical characterization of disordered protein liquid phases, The Use of Mass Spectrometry to Examine IDPs – Unique Insights and Caveats, Fluorescence Depolarization Kinetics to Study Conformational Preference, Structural Plasticity and Membrane Binding of Intrinsically Disordered Proteins, Characterizing the Function of Intrinsically Disordered Proteins in the Circadian Clock, and more. Breadth of experimental approaches and systems that will be covered The expertise of the contributors writing the articles

This open-end treatise on methods concerning protein separation had its beginning in an American Chemical Society symposium entitled'Con­ temporary Protein Separation Methods'which was held in Atlantic City, New Jersey in September 1974. The purpose of the symposium-and subse­ quently of the present work-was to review the available modern techniques and underlying principles for achieving one of the very important tasks of experimental biology, namely the separation and characterization of proteins present in complex biological mixtures. Physicochemical characterization was covered only as related to the parent method of fractionation and there­ fore involved mostly mass transport processes. Additionally, the presentation of methods for gaini. ng insight into complex interacting protein profiles was considered of paramount importance in the interpretation of separation patterns. Finally, specific categories of proteins (e. g., chemically modified, deriving from a specific tissue, conjugated to different moieties, etc.) require meticulous trial and selection andjor modification of existing methodology to carry out the desired separation. In such cases, the gained experience provides valuable guidelines for further experimentation. Although powerful techniques exist today for the separation and related physicochemical characterization of proteins, many biological fractionation problems require further innovations. It is hoped that the description in the present treatise of some of the available separation tools and their limitations will provide the necessary integrated background for new developments in this area.

Isoelectric focusing (IEF) is a high-resolution, stand-alone technique that can be used as an analytical method or tool for protein purification. The only current book on the market, the Handbook of Isoelectric Focusing and Proteomics is the ideal'one-stop'source for germane information in this discipline. This highly practical book also contains chapters on alternative methods that may pave the way in the search for efficient techniques for fractionating and purifying proteins. Complete with the history of IEF focusing to authors'insights and practical tips, this book is a must for anyone working in proteomics.• Is the only current book available on the subject • Includes author insights and practical tips• Is an ideal single source for students and researchers working in proteomics

Ultrafiltration is a pressure-driven, membrane-based separation process, which is used for a broad variety of applications, ranging from the processing of biological macromolecules to wastewater treatment. It has significant advantages over competing separation technologies. Food and biotechnological applications account for nearly 40% of the current total usage of ultrafiltration membranes. Protein bioseparation is an important component of this application segment. Ultrafiltration is used for protein concentration, desalting, clarification and fractionation (i.e. protein-protein separation). Concentration, desalting and clarification are technologically less demanding and have been in used in the bioprocess industry for some time. Protein fractionation, on the other hand, is a challenging proposition and is definitely a more recent development. This book focuses primarily on protein fractionation.

This publication details the isolation of proteins from biological materials, techniques for solid-liquid separation, concentration, crystallization, chromatography, scale-up, process monitoring, product formulation, and regulatory and commercial considerations in protein production. The authors discuss the release of protein from a biological host, selectivity in affinity chromatography, precipitation of proteins (both non-specific and specific), extraction for rapid protein isolation, adsorption as an initial step for the capture of proteins, scale-up and commercial production of recombinant proteins, and process monitoring in downstream processing.

It is a truism of science that the more fundamental the subject, the more universally applicable it is. Nevertheless, it is important to strike a level of “fundamentalness” appropriate to the task in hand. For -depth study of the mechanics of motor cars would tell one example, an in nothing about the dynamics of traffic. Traffic exists on a different “level” - it is dependent upon the existence of motor vehicles but the physics and mathematics of traffic can be adequately addressed by considering motor vehicles as mobile “blobs”,with no consideration of how they become mobile. To start a discourse on traffic with a consideration of the mechanics of motor vehicles would thus be inappropropriate. In writing this volume, I have wrestled with the question of the appropriate level at which to address the physics underlying many of the techniques used in protein isolation. I have tried to strike a level as would be used by a mechanic (with perhaps a slight leaning towards an engineer) - i.e. a practical level, offering appropriate insight but with minimal mathematics. Some people involved in biochemical research have a minimal grounding in chemistry and physics and so I have tried to keep it as simple as possible.

The book deals with the theory and practice of all electrophoretic steps leading to proteome analysis, i.e. isoelectric focusing (including immobilized pH gradients), sodium dodecyl sulphate electrophoresis (SADS-PAGE) and finally two-dimensional maps. It is a reasoned collection of all modern, relevant, up-to-date methodologies leading to successful fractionation, analysis and characterization of every polypeptide spot in 2-D map analysis. It includes chapters on the most sophisticated mass spectrometry developments and it helps the reader in navigating through the most important databases in proteome analysis, including step by step tours in selected sites. Yet, this book's unique strength and feature is the fact that it combines not only practice (in common with any other book on this topic) but also theory, by giving a detailed treatment on the most advanced theoretical treatments of steady-state techniques, such as isoelectric focusing and immobilized pH gradients. A lot of this theory is newly developed and presented to the public for the first time. Thus, this book should satisfy not only the needs of every day practitioners, but also the desires of the most advanced theoreticians in the field, who will surely appreciate the novel theories presented here. Also the methodological section contains several as yet unpublished protocols, correcting some of the existing ones and showing the pitfall and limitations of even well ingrained protocols in proteome analysis, which are here critically re-evaluated for the first time.

This book covers the fundamentals of protein inactivation during bioseparation and the effect on protein processing. Bioseparation of Proteins is unique because it provides a background of the bioseparation processes, and it is the first book available to emphasize the influence of the different bioseparation processes on protein inactivation.Bioseparation of Proteins covers the extent, mechanisms of, and control of protein inactivation during these processes along with the subsequent and essential validation of these processes. The book focuses on the avoidance of protein (biologicalproduct) inactivation at each step in a bioprocess. It compares protein inactivation exhibited during the different bioseparation processes by different workers and provides a valuable framework for workers in different areas interested in bioseparations.Topics include separation and detection methods; estimates of protein inactivation and an analysis of this problem for different separation processes; strategies for avoiding inactivation; the molecular basis of surface activity and protein adsorption,process monitoring, and product validation techniques; and the economics of various bioseparation processes and quality control procedures.Key Features• Protein inactivation and other aspects of biological stability are critical to an effective bioseparation process; This book is a detailed and critical review of the available literature in an area that is essential to the effectiveness, validation, and economics of bioseparation processes for drugs and other biological products; Conveniently assembled under one cover, the survey of the literature and resulting perspective will greatly assist engineers and chemists in designingand improving their own processes; Key features of the text include:• detailed data on biological stability under various bioseparation conditions• extensive case studies from the literature on separation processes, validation, and economics• simplified analysis of protein refolding and inactivation mechanisms• consideration of adsorption theories and the effect of heterogeneity• coverage of both classical and novel bioseparation techniques, including chromatographic procedures

This book enables the novice to understand the'whys'and'hows'of electrophoresis and to initiate and complete an electrophoretic investigation from beginning laboratory organization to publishing results.