Immobilized Affinity Ligand Techniques

This book is a practical guide to the preparation and use of immobilized affinity ligands for purification, catalysis, and analysis. Special emphasis is given to immunochemical techniques including antibody isolation, preparation of antibody fragments using immobilized enzymes, and immunoaffinity chromatography. The book provides easy-to-follow, well-tested protocols to allow the uninitiated to use these techniques to the maximum advantage with minimum hassle. In addition, it shows researchers how to save money by making their own optimized affinity supports. Matrix activation: Ligand immobilization, Binding and elution of target molecules, Enzyme catalysis on solid supports, Analytical affinity chromatography, Isolation/purification of antibodies, Preparation of antibody fragments, Immunoaffinity chromatography, Immobilization of nucleic acids, Use of immobilized ligands for removal of trace contaminants Practical advice on choosing: Matrices, Spacers, Methods of activation and coupling Background information and insights on: Affinity interactions, The ease and power of affinity chromatography, Attaching molecules to insoluble supports, Matrices currently in use, Over 20 methods of activation, Spacers, Extensive References

A timely, accessible survey of the multidisciplinary field of bioanalytical chemistry Provides an all in one approach for both beginners and experts, from a broad range of backgrounds, covering introductions, theory, advanced concepts and diverse applications for each method Each chapter progresses from basic concepts to applications involving real samples Includes three new chapters on Biomimetic Materials, Lab-on-Chip, and Analytical Methods Contains end-of-chapter problems and an appendix with selected answers

Affinity chromatography, with its exquisite specificity, is based upon molecular recognition. It is a powerful tool for the purification of biomolecules. In recent years, numerous new applications and modified techniques have been derived from gro- specific interactions and biological recognition principles. An up-to-date review of the past, current, and future applications of affinity chromatography has been presented in the introductory chapter by Meir Wilchek and Irwin Chaiken. Though many of these new applications and techniques are well documented in the literature, it is often difficult to find methods that are written with the intent of helping new practitioners of affinity chromatography. This volume on Affinity Chromatography: Methods and Protocols is intended for the novice, as well as for - perts in the field. The protocols are written by experts who have developed and/or successfully employed these methods in their laboratories. Each chapter describes a specific technique, and since the book is intended to help the beginner, each technique is described simply and clearly, making sure that all relevant steps are included, assuming no previous knowledge. Each chapter contains an introduction describing the principles involved, followed by a Materials and Methods section, which lays the groundwork for the reader to conduct experiments step-by-step, in an orderly fashion. The following Notes section, which describes many of the problems that can occur, makes suggestions for overcoming them, and provides alternate procedures. These are precisely the sort of important, practical details that never seem to appear in the published literature.

Bioaffinity chromatography has increasingly become the method of choice for the purification, determination or removal of many biologically active substances.

Bioconjugate Techniques, 2nd Edition, is the essential guide to the modification and cross linking of biomolecules for use in research, diagnostics, and therapeutics. It provides highly detailed information on the chemistry, reagent systems, and practical applications for creating labeled or conjugate molecules. It also describes dozens of reactions with details on hundreds of commercially available reagents and the use of these reagents for modifying or cross linking peptides and proteins, sugars and polysaccharides, nucleic acids and oligonucleotides, lipids, and synthetic polymers. A one-stop source for proven methods and protocols for synthesizing bioconjugates in the lab Step-by-step presentation makes the book an ideal source for researchers who are less familiar with the synthesis of bioconjugates More than 600 figures that visually describe the complex reactions associated with the synthesis of bioconjugates Includes entirely new chapters on the latest areas in the field of bioconjugation as follows: Microparticles and nanoparticlesSilane coupling agentsDendrimers and dendronsChemoselective ligationQuantum dotsLanthanide chelatesCyanine dyesDiscrete PEG compoundsBuckyballs,fullerenes, and carbon nanotubesMass tags and isotope tagsBioconjugation in the study of protein interactions

Affinity chromatography is a type of liquid chromatography that uses a biologically related agent as a stationary phase to purify or analyze specific sample components. This chapter examines the basic components and principles of affinity chromatography and various applications of this method. The general types of ligands, immobilization schemes, support materials, and elution conditions that can be employed in affinity chromatography are first examined. Various methods in this field are then considered, including bioaffinity chromatography, immunoaffinity chromatography, dye-ligand and biomimetic affinity chromatography, immobilized metal-ion affinity chromatography, and analytical affinity chromatography. The types of ligands utilized in each of these methods are described, along with common applications of these techniques. Several related developments are also discussed, such as the use of affinity chromatography with mass spectrometry, miniaturized systems, molecularly imprinted polymers, and aptamers.

Chapter 1: Principles on membrane and membrane processes -- Chapter 2: Ultrafiltration -- Chapter 3: Microfiltration -- Chapter 4: Virus Filtration -- Chapter 5: Membrane chromatography -- Chapter 6: Membranes for the Preparation of Emulsions and Particles -- Chapter 7: Other Membrane Processes -- Chapter 8: Some Perspectives.

Bioaffinity chromatography is now the preferred choice for the purification, determination or removal of many biologically active substances. The book includes information on biologically active substances with their affinants, solid supports and methods of coupling, summarized in tables covering classical, high-performance liquid and large-scale bioaffinity chromatography. Optimization of the preparation and the use of highly active and stable biospecific adsorbents is discussed in several chapters. Following a chapter dealing with the choice of affinity ligands, affinity-sorbent bonding is described in detail. Other chapters give information on solid supports, the most common coupling procedures and a general discussion of sorption and elution. Several applications of bioaffinity chromatography are described, e.g. quantitative evaluation of biospecific complexes and many applications in medicine and in the biotechnology industry.