© 1999 • 495 pp., illus., appendices, index
Concealed wire binding • $159 • ISBN  978-087969544-6

This title also available in: Cloth

Few technical manuals have become standards in biomedicine. Antibodies: A Laboratory Manual, by Ed Harlow and David Lane, has had that status for a decade. Now there is a new and even higher standard—Using Antibodies: A Laboratory Manual.

Harlow and Lane have completely revised their guide to the use of immunoglobulin reagents in the laboratory. Chapters have been entirely rewritten, reorganized, and updated to provide background, context, and step-by-step instructions for techniques that range from choosing the right antibody and handling it correctly, to the proper methods for characterizing antigens in cells and solutions. New chapters on tagging proteins and epitope mapping are included.

Rather than presenting an array of solutions for working with antibodies and antigens, Using Antibodies instead identifies in each case the best approach to specific problems. These recommendations include more detail in the protocols, extensive advice on avoiding and solving problems, information regarding proper controls, and extensive illustration of theory, methods, and results, both good and bad. An additional bonus included with this manual is a set of Portable Protocols, step-by-step instructions for the most frequently used and essential techniques printed on spill-proof, durable cards that can be annotated and used directly at the bench.

The expert advice in Using Antibodies is presented using an imaginative design with extensive use of color and graphic elements calculated to help readers plan and execute their experiments efficiently and accurately. A newly available type of binding will maintain the manual's integrity during years of use.

This new manual reflects a decade's additional research experience by two outstanding scientists of international reputation. Since writing the previous manual, Ed Harlow has received many awards, notably the General Motors and Bristol Myers prizes for cancer research, and he was elected to the National Academy of Sciences. David Lane, also the winner of many awards, such as the Yvette Mayent Prize and the Paul Ehrlich and Ludwig Darmstaedter Award, was elected as a Fellow of the Royal Society. The over-the-shoulder advice these experts provide in Using Antibodies will lead all laboratory investigators to success in using these techniques, regardless of experience. Using Antibodies is a required resource for every laboratory in which genes, cells, and proteins are studied.

  "For nearly a decade, this book has been the standard resource in immunological research. The authors have now completely revised the manual, incorporating both praise and criticism from frequent users of the previous version, as well as from world-renowned experts.

The first major change is the separation of information into two volumes. Using Antibodies: A Laboratory Manual covers methods for using antibodies to staining cells and tissues, immunoprecipitation, immunoblotting, immunoaffinity purification, tagging proteins and epitope mapping. The second volume, Making Antibodies, will be published separately. This was a wise move, considering the mass of information presented by the authors, and it allows them to provide adequate background information on selected procedures, antibody structure and function, choosing and handling antibodies, and antigen-antibody interactions.

The protocols are detailed, clear, concise and easy to follow. The authors also outline their reasons why certain techniques are preferred over others. The sections on troubleshooting and common problems encountered with use of the protocols are invaluable. Another key change lies in the construction of the manual. The previous comb-bound edition was not sturdy enough to withstand repeated use in the laboratory, but the current edition features a hardcover bound with concealed wire, so that it can lie flat and stand up to heavy use. In addition, nine laminated ‘Portable Protocol’ sheets can be used directly at the bench … .

Using Antibodies: A Laboratory Manual is an outstanding revision of a classic work that is destined to become the new standard resource for antibody techniques used in immunological research."
      —The Quarterly Review of Biology

  "…this book lives up to its expectation: it is easy to follow with many tables listing advantages and disadvantages of most protocols, how-to illustrations, best approach protocols for beginners to get started, and illustrations of good and bad results. This manual is a ‘must have’ for laboratories routinely using antibodies."
      —In Vitro Cellular & Developmental Biology - Animal

Preface

Dedication

Chapter 1: Antibody Structure And Function

Chapter 2: Antibody–Antigen Interactions

Chapter 3: Choosing Antibodies

Chapter 4: Handling Antibodies

Storing Antibodies

Purifying Antibodies

Labeling Antibodies

Chapter 5: Staining Cells

Major Constraints

Choosing The Correct Antibody

Protocols For Immunostaining Of Tissue Culture Cells

Variations On The Immunostaining Procedure

Chapter 6: Staining Tissues

Major Constraints

Choosing the Correct Antibody

Immunostaining Tissues With Polyclonal Or Monoclonal Antibodies

Protocols For Staining Tissue Sections

Variations For Staining Yeast

Variations For Staining Drosophila

Protocols For Antigen Retrieval

Chapter 7: Immunoprecipitation

Major Constraints

Choosing The Correct Antibody

Immunoprecipitation Protocols

Variations On The Immunoprecipitation Procedure

Chapter 8: Immunoblotting

Major Constraints

Choosing The Correct Antibody

Immunoblotting Protocols

Variations In The Detection Methods

Variations In The Immunoblotting Technique

Chapter 9: Immunoaffinity Purification

Major Constraints

Choosing The Correct Antibody

Immunoaffinity Purification With Polyclonal And Monoclonal Antibodies

Protocols For Immunoaffinity Purification

Variations On Immunoaffinity Purification

Chapter 10: Tagging Proteins

Major Constraints

Why Tag Your Protein

Tags For Protein Detection

Tags For Protein Purification

Choosing The Right Tag

Deciding Where To Position The Tag

How To Tag Your Protein

Chapter 11: Epitope Mapping

Determining The Structural Requirements Of An Epitope

Choosing An Epitope-mapping Method

Mapping By Competition Assay

Mapping By Expression Of Gene Fragments

Mapping Using Synthetic Peptides

Synthesis Or Purchase Of The Peptide Set

Alternate Approaches To Epitope Mapping

Appendix I: Electrophoresis

Appendix II: Protein Techniques

Appendix III: General Information

Appendix IV: Cautions

Appendix V: Trademarks

Appendix VI: Suppliers