Capillary Gel Electrophoresis and Related Microseparation Techniques book cover

Capillary Gel Electrophoresis and Related Microseparation Techniques

This book has been designed to cover all theoretical and practical aspects of capillary gel electrophoresis. It also provides an excellent overview of the key application areas of nucleic acid, protein and complex carbohydrate analysis, affinity based methodologies, micropreparative aspects, as well as related microseparation methods. It will also give the reader a better understanding of how to utilize this technology, and determine which actual method will provide appropriate technical solutions to problems that the reader may have perceived as being more fundamental. This book can also serve as a textbook for undergraduate and graduate courses in analytical chemistry, analytical biochemistry, molecular biology and biotechnology courses.

Audience
Academics, industrial analytical chemists, regulatory agencies

Hardbound, 350 Pages

Published: May 2014

Imprint: Elsevier

ISBN: 978-0-444-52234-4

Contents

  • PrefaceList of SymbolsAbbreviations1. INTRODUCTION 1.1. History of gel electrophoresis1.2. Modes of capillary gel electrophoresis1.3. Basic principles and theory of capillary gel electrophoresis1.3.1. Electrophoretic Migration1.3.2. Secondary Equilibrium1.3.3. Efficiency and Resolution1.3.4. Capillary gel electrophoresis of DNA1.4. Capillary electrophoresis in anticonvective media1.5. Band broadening in capillary gel electrophoresis1.6. Power dissipation1.7. Comparison with slab gel electrophoresis1.8. References2. SEPARATION MATRIX AND COLUMN TECHNOLOGY2.1. Capillary Coatings2.1.1. Covalent coatings2.1.2. Physical (non-covalent) surface coverage2.2. Separation Matrices2.2.1. Cross-linked (chemical) gels, structure and properties2.2.2. Non cross-linked linear polymers (physical gels)2.2.2.1. Non-cross-linked polyacrylamide and agarose2.2.2.2. Derivatized celluloses2.2.2.3. Polyethylene oxide and polyvinylpyrrolidone2.2.3. Copolymers, composite gels and other alternative matrices2.3. Techniques for Preparing Gel Filled Capillaries2.3.1. Typical gel formulations2.3.2. Choosing the appropriate separation media2.4. Alternative Matrices (composite gels, pluronics, sol-gel systems, block polymers)2.5. References3. INSTRUMENTATION 3.1. Sample Introduction Methods3.1.1. Electrokinetic injection into high viscosity gels3.1.2. Pressure and vacuum injection into low viscosity polymer solutions3.1.3. Sample stacking3.1.4. Effect of sample overloading3.1.5. Injection related artifacts3.2. Detection Systems 3.2.1. Requirements of capillary gel electrophoresis detectors3.2.2. UV absorbance and diode array detection3.2.3. Laser induced fluorescence (LIF) detection3.2.4. Indirect detection methods3.2.5. Electrochemical detection3.2.6. Radioisotope detectors and other miscellaneous detection methods3.2.7. Coupling to Mass Spectrometers3.3. Operation Variables3.3.1. Gel concentration, Ferguson plots3.3.2. Temperature and voltages, Arrhenius plots3.3.3. Capillary dimensions3.3.4. Buffer systems3.3.5. Organic modifiers, non-aqueous electrophoresis3.3.6. Complexing additives3.4. References4. APPLICATIONS4.1. Capillary Gel Electrophoresis of DNA4.1.1. Sample purification, injection and separation4.1.2. Single stranded oligonucleotides and DNA sequencing4.1.3. ds DNA fragments and PCR products4.1.4. RNA analysis4.1.5. Large chromosomal DNA4.1.6. Antisense DNA and phosphodiester oligonucleotides4.1.7. Biomedical and forensic applications4.2. Capillary Gel Electrophoresis of Proteins4.2.1. Fundamentals of capillary SDS gel electrophoresis4.2.2. The Ferguson method4.2.3. Fluorescent labeling techniques4.2.4. Ultrafast separations4.2.5. Capillary isoelectric focusing4.2.6. Applications in biotechnology 4.3. Capillary Gel Electrophoresis of Carbohydrates4.3.1. Analytical glycobiology4.3.2. Release of N and O linked oligosaccharides from glycoproteins4.3.3. Fluorophore labeling reactions4.3.4. Capillary gel electrophoresis separation of labeled glycans4.3.5. Selected applications4.4. Capillary Affinity Gel Electrophoresis4.4.1. Principles of capillary affinity gel electrophoresis4.4.2. Effect of affinity agent concentration and pH 4.4.3. Chiral additives, types of chiral selectors4.4.4. Capillary affinity gel electrophoresis of DNA 4.4.5. Capillary affinity gel electrophoresis of proteins4.4.6. Capillary affinity gel electrophoresis of enantiomers4.4.7. Pharmaceutical applications4.5. Other, Miscellaneous Applications4.6. References5. MICROPREPARATIVE CAPILLARY GEL ELECTROPHORESIS5.1. Theoretical background of micropreparative applications5.2. High-resolution separations5.3. Field programming for micropreparative fraction collection5.4. Rapid and large scale expressed sequence tag (EST) generation5.5. High throughput cloning5.6. References6. RELATED MICROSEPARATION TECHNIQUES6.1. Ultra-Thin-Layer Gel Electrophoresis6.1.1. Early miniaturization attempts6.1.2. Polyacrylamide gel based systems, DNA sequencing6.1.3. Automated ultra-thin-layer gel electrophoresis using agarose gels6.1.4. High throughput analysis of dsDNA fragments and proteins6.1.5. Two dimensional approaches6.1.6. Biomedical applications6.2. Gel Electrophoresis in Microfabricated Devices6.2.1. Modular system design6.2.2. Microfabrication techniques6.2.3. Fluid manipulation and injection6.2.4. Gel electrophoresis in microchips6.2.5. Analysis of single and double stranded DNA6.2.6. Analysis of proteins6.2.7. Integrated microfabricated biodevices6.3. References7. APPENDIX. Addresses of some suppliers of instrumentation, chemicals and reagents8. GLOSSARY9. SUBJECT INDEX

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