Over the past thirty years, the development of the Western blot has revolutionized the fields of biomedical research and medical diagnostics. In this volume, expert researchers present numerous techniques based on the Western blot, providing detailed, readily reproducible methods, tips and alternatives directly and easily transferable to the laboratory setting.
Chapters offer a large number of variations on the theme of protein transfer to solid support followed by detection, presenting both adaptations of traditional techniques as well as completely original methods of protein blotting.
Protein Blotting and Detection: Methods and Protocols Editors Biji T. Kurien, PhD and R. Hal Scofield, MD Table of Contents Preface- R. Hal Scofield and Biji T. Kurien Contributors 1. Origins of protein blotting- Dr. Harry Towbin 2. Western Blotting: Remembrance of Things Past-W. Neal Burnette 3. Introduction to protein blotting- Biji T. Kurien and R. Hal Scofield BLOTTING BY DIFFUSION 4. Simultaneous Immunoblotting Analysis with Activity Gel Electrophoresis and 2-D Gel Electrophoresis- Der-Yen Lee and Geen-Dong Chang 5. Diffusion blotting for rapid production of multiple identical imprints from sodium dodecyl sulfate polyacrylamide gel electrophoresis on a solid support- Ingrid Olsen and Harald G. Wiker 6. Tryptic peptide purification using polyvinylidene difluoride membrane for matrix assisted laser desorption ionization time of flight mass spectrometry -Biji T. Kurien and R. Hal Scofield 7. Affinity immunoblotting for analysis of antibody clonotype distribution in a lupus patient developing anti-Ro 60 over time-Biji T. Kurien and R. Hal Scofield 8. Non-electrophoretic bi-directional transfer of a single SDS-PAGE gel with multiple antigens to obtain twelve immunoblots - Biji T. Kurien and R. Hal Scofield 9. Slice blotting-Graeme Lowe 10. Localizing Proteins by Tissue Printing - Rafael Pont-Lezica 11. Dot-Immunobinding Assay (Dot-Iba) - Sumi Surendran, Annamma Mathai and Vishnampet Venkataraman Radhakrishnan 12. Capillary Blotting of Glycosaminoglycans on Nitrocellulose Membranes after Agarose-gel Electrophoresis Separation- Nicola Volpi and Francesca Maccari MODIFICATIONS OF ELECTROBLOTTING 13. Quantitative computerized western blotting- Dalit Talmi-Frank, Charles L. Jaffe and Gad Baneth 14. Cationic electrophoresis and eastern blotting - Engelbert Buxbaum 15. A miniaturized blotting system for simultaneous detecting of different autoantibodies - Ulrich Canzler, Holger Bartsch, Kai Grossmann, WernerLehmann, Karsten Conrad, Biji T. Kurien, Yaser Dorri, R. Hal Scofield, Michael P. Bachmann 16. Transfer and multiplex immunoblotting of a paraffin embedded tissue- Joon-Yong Chung and Stephen M. Hewitt 17. Multistrip Western Blotting to increase quantitative data output- Anatoly Kiyatkin and Edita Aksamitiene 18. The Western blotting/Edman sequencing technique using polyvinylidene difluoride (PVDF) membrane: Setsuko Komatsu 19. Blotting from PhastGel to membranes by ultrasound- Joseph Kost 20. Ultra-rapid electrophoretic transfer of high and low molecular weight proteins using heat- Biji T. Kurien and R. Hal Scofield 21. Strip immunoblotting of multiple antigenic peptides to nitrocellulose membrane- Biji T. Kurien and R. Hal Scofield 22. Affinity purification of autoantibodies from an antigen strip excised from a nitrocellulose protein blot - Biji T. Kurien and R. Hal Scofield 23. Double-blotting: A Solution to the Problem of Non-specific Binding of Secondary Antibodies in Immunoblotting Procedures - Francoise Lasne 24. Efficient electroblotting of very large proteins using a vertical agarose electrophoresis system- Marion L. Greaser and Chad M. Warren 25. Semi-dry Protein Transfer and Immunodetection of P-selectin Using an Antibody to its C-terminal Tag- Padmaja Mehta 26. The Multiple Antigen Blot Assay-MABA, a simple, versatile and multi-purpose immunoenzymatic technique- Oscar O Noya, Sandra Losada, Marilyan Toledo, and Belkisyole Alarcon de Noya 27. Blotting from immobilized pH gradient gels: Application to total cell lysates - Harry Towbin 28. Immunoprecipitation-Western blot for proteins of low abundance - Edward Trieu, Jody J. Gross and Ira Targoff 29. Native electrophoresis and Western blot analysis (NEWeB): Method and Applications- Ioannis N. Manoussopoulos and Mina Tsagris 30. Grid-immunoblotting- Olga Yeliosof and R. Hal Scofield MANIPULATIONS OF ELECTROBLOTTED MEMBRANES 31. Sequential Use of Immunoblots for Characterization of Autoantibody Specificities-Holger Bartsch, Michael Bachmann 32. Renaturation of transferred recombinant protein on PVDF by calcium- Biji T. Kurien, Michael Bachmann, R. Hal Scofield 33. Intermittent Microwave Irradiation Facilitates Antigen- Antibody Reaction in Western Blot Analysis- Yu-Ting Liu and Shinya Toyokuni 34. Detection of Protein-Protein Interactions by Far-Western Blotting- Bruce J. Meyer and Kazuya Machida 35. On-membrane tryptic digestion of proteins for mass spectrometry analysis- Jose L. Luque-Garcia and Thomas A. Neubert 36. Western Blot Analysis of Adhesive Interactions under Fluid Shear Conditions: The Blot Rolling Assay - Robert Sackstein and Robert Fuhlbrigge OVERVIEW CHAPTERS 37. ELISPOT assay on membrane microplates-Alexander E. Kalyuzhny 38. A Brief Review of Other Notable Protein Blotting Methods -Biji T. Kurien and R. Hal Scofield ANTIGEN DETECTION 39. Phosphoprotein Detection on Protein Electroblot Using a Phosphate-Specific Fluorophore - Lee B. Bockus and R. Hal Scofield 40. Rapid, antibody-free detection of recombinant proteins on blots using enzyme fragment complementation- Neil W. Charter, Joe Horecka, Chin-Yee Loh, Albert Doan, Tom Wehrman and Keith R. Olson. 41. Fluorescent Labeling of Proteins and Its Application to SDS-PAGE and Western Blotting- F. Javier Alba, Salvador Bartolome, Antonio Bermudez, and Joan-Ramon Daban 42. Whole and Strip Nitrocellulose Membrane as well as a New-Line-Immunoblotting of Antigen Using the Chemiluminescence Technique -Yaser Dorri, Ali Khalili and R. Hal Scofield 43. Protein Stains to Detect Antigen on Membranes - Anil D'Souza and R. Hal Scofield 44. Use of non-radioactive detection method for North- and South-Western Blot- Claudia Franke, Daniel Grafe, Holger Bartsch, Michael Bachmann 45. Immunoblotting using Radiolabeled Reagents for Detection- Holger Bartsch Claudia Franke, Michael Bachmann 46. Detection of Protein Carbonyls by Means of Biotin Hydrazide --Streptavidin Affinity Methods- Kenneth Hensley 47. "Rainbow" Western Blotting- Stanislaw Krajewski 48. Multiple antigen detection (MAD) western blotting- Stanislaw Krajewski, Xianshu Huang and Maryla Krajewska 49. Detection of calcium binding by Ro 60 multiple antigenic peptides on nitrocellulose membrane using Quin-2- Biji T Kurien, Michael Bachmann, R. Hal Scofield 50. Detection of La/SS-B by Western Blot using Nanogold-Tagged Antibodies and Silver Enhancement- Shannon Maier, Sherry Hubbel and R. Hal Scofield 51. Imaging Systems for Westerns: Chemiluminescence Vs. Infrared Detection- Suresh T. Mathews, Eric P. Plaisance, and Teayoun Kim 52. A fluorescent co-detection system for Immunoblotting and Proteomics through ECL-Plex and CyDye labeling- Ciara A. McManus, Pamela M. Donoghue, and Michael J. Dunn 53. Visualization of unstained protein bands on PVDF -- Jun Park, Masaharu Mabuchi and Ajay Sharma 54. Improved Protein Detection Using Cold Microwave Technology- Aaron G. Smith, Jyothi Jayaram, Carol B. Johnson, E. Ann Ellis, Stanislav Vitha, Ellen W. Collisson and Andreas Holzenburg 55. TLC Blot (Far-eastern Blot) and Its Applications- Takao Taki, Naoko Goto-Inoue, Takahiro Hayasaka, Mitsutoshi Setou OVERVIEW CHAPTER 56. A brief review of other notable protein detection methods on blots -Biji T. Kurien and R. Hal Scofield CONCLUDING CHAPTER 57. Sending secret messages on nitrocellulose membrane and the use of a molecular pen for orientation marking in ECL membrane assays- Biji T. Kurien and R. Hal Scofield
From the reviews: "This informative, hands-on-guide is essential for all investigators interested in learning techniques based on Western blotting which they can easily transfer to their laboratories. ! The book is intended for any investigator interested in learning a new technique based on Western blotting. ! this hands-on guide will be quite useful and helpful for both investigators and students working in these areas." (Omer Iqbal, Doody's Review Service, October, 2009)