Microwave Assisted Proteomics - Hardcover

Lill, Jennie R

Hardcover

ISBN 10: 085404194X ISBN 13: 9780854041947

Verlag: Royal Society of Chemistry, 2009

Inhaltsangabe

This book is an introduction to microwave-assisted bio-analytical methods with an emphasis on sample preparation for proteomic analyses. Its objective is to provide a history and background for the methods described. The book summarizes the development and application of these protocols from both the literature in this field as well as from the experiences of a variety of laboratories conducting microwave-assisted protocols. In addition to describing the evolution of the microwave-assisted methodologies, the book also offers a concise protocol section with practical advice on how to implement these methods in any laboratory. Microwave-assisted proteomics is a rapidly evolving field and this book captures the main areas that fall under this emerging arena.

Über diesen Titel

Über die Autorin bzw. den Autor

Jennie R Lill, Genentech Inc, San Francisco, USA has spent 11 years in proteomic mass spectrometry research. She has a B.Sc. in Biological Sciences from Warwick University, and an M.Sc. in Biotechnology from Nottingham Trent University and a Ph.D. Thesis entitled "Characterisation of MHC class I peptides by reverse phase chromatography and ion trap mass spectrometry" from Nottingham Trent University.

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This book is an introduction to microwave-assisted bio-analytical methods with an emphasis on sample preparation for proteomic analyses. Its objective is to provide a history and background for the methods described and the book summarizes the development and application of these protocols from both the literature in this field as well as from the experiences of a variety of laboratories conducting microwave-assisted protocols. In addition to describing the evolution of the microwave-assisted methodologies, it includes mechanisms of action as well as an overview of the types of instrumentation available to help the reader develop the best experimental design for their own budget and experimental aims. The key features of this book: * An overview of the history of microwave-assisted proteomics * An in-depth discussion on several microwave-assisted methods including - enzymatic and chemical proteolytic reactions, microwave-assisted acid hydrolysis for N-terminal sequencing and for amino acid analysis * A discussion of the mechanisms involved * An overview of instrumentation types and limitations * A concise and practical protocol section for the researcher to carry out the experiments in their own laboratory Microwave-assisted proteomics is a rapidly evolving field and this book captures the main areas that fall under this emerging arena.

Auszug. © Genehmigter Nachdruck. Alle Rechte vorbehalten.

Microwave-Assisted Proteomics

By Jennie Rebecca Lill

The Royal Society of Chemistry

Copyright © 2009 Jennie Rebecca Lill
All rights reserved.

ISBN: 978-0-85404-194-7

Contents

Acknowledgements, xi, 
Chapter 1 Evolution of Microwave Irradiation and Its Introduction to the Biosciences, 1, 
Chapter 2 Microwave Instrumentation in the Biosciences, 10, 
Chapter 3 Mechanisms of Microwave-Assisted Action, 21, 
Chapter 4 Microwave-Assisted Enzymatic Digestions, 28, 
Chapter 5 Microwave-Assisted Chemical Digestion of Proteins, 43, 
Chapter 6 Microwave-Assisted Acid Hydrolysis, 56, 
Chapter 7 Microwave-Assisted Discovery and Characterization of Posttranslational Modifications, 71, 
Chapter 8 Recent Microwave-Assisted Applications in the Life Sciences, 92, 
Chapter 9 Epilogue: To Microwave or Not To Microwave?, 110, 
Chapter 10 Microwave-Assisted Proteomic Protocols, 112, 
Subject Index, 119, 

CHAPTER 1

Evolution of Microwave Irradiation and Its Introduction to the Biosciences

Abstract

Since the conceptualization of the electromagnetic spectrum, through the development of the magnetron microwave energy has been utilized in many aspects and disciplines of science. Although adopted by many industries over the past quarter of a century, it is only within the past few years that microwave irradiation has been evaluated as a useful tool in the biochemical and chemical preparation of proteins and other biomolecules. This chapter describes the evolution of the magnetron and some early applications of microwave assistance in the bioanalytical sciences.

1.1 Microwave Radiation

The electromagnetic spectrum is a continuum of all electromagnetic waves arranged according to frequency and wavelength. Microwaves occupy the electromagnetic spectrum between infrared radiation and radio waves and have wavelengths between 0.01 and 1 m with a frequency range between 0.3 and 30 GHz (Figure 1.1). Most commercially available microwaves have a narrower range at around 2.5 GHz. Microwave energy is a natural phenomenon which can be induced when electric current flows through a conductor, for example an antenna, a transmitter chip or a magnetron.

1.2 History and Evolution of the Magnetron

The theory of microwave irradiation was first documented in 1864 by James Clerk Maxwell, a Scottish mathematician and theoretical physicist. In the eponymous Maxwell's equations he described a unified model for electro-magnetism and paved the way for modern physics. It was not until over twenty years later that microwave irradiation was physically demonstrated by Heinrich Hertz. Hertz was a German physicist who demonstrated the existence of electromagnetic radiation by building antennae and apparatus to produce and detect high-frequency radio waves. Microwaves were produced using various apparatus up until the 1920s when Albert Hull, a researcher at General Electric's research laboratories, invented the simple two-pole magnetron, or split-anode magnetron. Although revolutionary at its time, the two-pole magnetron was relatively inefficient and was soon superseded by the resonant-cavity magnetron which proved to be more efficient and convenient.

The first half of the twentieth century become synonymous with large-scale war and scientific innovation, the combination of which led to the development of many technologies including the introduction of radar. In a deal between British and American researchers, the cavity magnetron was developed into a viable radar system, and by 1941 magnetrons for radar systems were being manufactured at a rate of 17 per day at Raytheon. It was during this time that a researcher at Raytheon, Percy Lebaron Spencer, made two important discoveries. Firstly, he was awarded the Distinguished Public Service Award by the US Navy for significantly improving the manufacturing process of magnetrons and increasing production more than 100-fold. Secondly, perhaps more famously, in 1945 while standing in front of an open magnetron he noticed that a chocolate bar had melted in his pocket. After several other "tests" including popping popcorn and exploding eggs he concluded that microwave radiation could be tailored for use in cooking devices and hence the invention of the microwave oven.

By 1947 the first commercial microwave oven had been manufactured by Raytheon, although during the first few years of commercialization these ovens stood at nearly 6 feet tall and weighed over 700 pounds. By the 1970s micro- wave ovens had become much more accommodating for household use, and by 1975 the sales of microwave ovens started to exceed those of gas oven ranges in the USA.

In 1978 the first commercial microwave for laboratory use was introduced by CEM and, since then, laboratory microwaves have increased in sophistication and utility to include models specific for the chemical and biological sciences. Figure 1.2 shows the time scale of the evolution of the magnetron.

1.3 Microwaves as a Catalysis Tool in Organic and Inorganic Chemistry

In 1986 the first reports of high-speed chemical synthesis with microwave assistance were published. Since then there have been numerous publications describing microwave-assisted synthetic reactions where most researchers observe shorter reaction times, increased yields and cleaner syntheses due to reduced by-product formation or side reactions (for a comprehensive review, see Alcazar et al. ). Originally, chemists would employ microwave assistance only for those reactions that proved troublesome or that resulted in poor yields. Nowadays, however, as instrumentation and an understanding of the mechanisms involved have matured, chemists routinely employ microwave-catalyzed protocols at the first stage of method development. Indeed for reactions involving highly polar reagents or metal catalysis, microwave irradiation is confirmed as the most valuable mode of heating available, and standard protocols for polymer synthesis and process control typically employ microwave assistance.

1.4 Microwave-Assisted Staining of SDS-PAGE and PVDF Membrane-Embedded Proteins

One of the first microwave-assisted applications in the biological setting was the fixing, staining and destaining of sodium dodecyl sulfate polyacrylamide gels and poly(vinylidine difluoride) (PVDF) membranes. Sodium dodecyl sulfate poly-acrylamide gel electrophoresis (SDS-PAGE) is a highly valuable technique employed in biochemistry, genetics and molecular biology to separate biomolecules according to their electrophoretic mobility. In the separation of proteins, the electrophoretic mobility is dictated by the length of the polypeptide chain or molecular weight as well as higher order protein folding, posttranslational modifications and other factors. After proteins are separated on the gel, the sample is typically fixed with a reagent to mobilize the gel and to stop migration or dispersion. Fixation is typically performed in a high percentage of methanol, which can clean up the gel from any remaining material from the SDS running buffer. After fixation the gel is stained using one of the many stains available, for example Coomassie Blue or silver stain. Traditional staining protocols recommend immersion of the gel or membrane into stain solution for many hours (often an over night incubation). After staining, gels are destained to remove background stain, and to allow the bands corresponding to the proteins of interest to be visualized. Proteins may also be electro blotted from the gel onto PVDF membranes whereby the sample is more...

Details zur Ausgabe

• Verlag: Royal Society of Chemistry
• Erscheinungsdatum: 2009
• Sprache: Englisch
• ISBN 10: 085404194X
• ISBN 13: 9780854041947
• Einband: Gebundene Ausgabe
• Anzahl der Seiten: 138
• Kontakt zum Hersteller: Nicht verfügbar
• Verantwortliche Person: gpsr@libri.de; gpsr@libri.de; ; Friedensallee 273; Hamburg; 22763; Deutschland