Epstein-Barr Virus Protocols / Edition 1

Epstein-Barr Virus Protocols / Edition 1

by Joanna B. Wilson
     
 

Joanna Wilson and Gerhard May have assembled a collection of the key m olecular biology protocols used in the analysis of Epstein-Barr virus (EBV), along with a series of valuable immunology, cell biology, and t ransgenic mouse protocols. These readily reproducible techniques inclu de methods for gene expression with mini-EBV plasmids, for expression analysis by

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Overview

Joanna Wilson and Gerhard May have assembled a collection of the key m olecular biology protocols used in the analysis of Epstein-Barr virus (EBV), along with a series of valuable immunology, cell biology, and t ransgenic mouse protocols. These readily reproducible techniques inclu de methods for gene expression with mini-EBV plasmids, for expression analysis by FISH, for EBV detection and quantitation, and for cell pro liferation and death assays. In addition, there are EBV-based vectors, an up-to-date map of EBV, a comprehensive table of available latent p rotein antisera, and assays from in vitro to cell to organ to organism levels. Timely and highly practical, Epstein-Barr Virus Protocols pro vides powerful tools for elucidating the life cycle of EBV and its hos t interactions, work that promises the emergence of major new treatmen ts and cures for EBV associated diseases, including several forms of h uman cancer.

Product Details

ISBN-13:
9780896036901
Publisher:
Springer-Verlag New York, LLC
Publication date:
03/23/2001
Series:
Methods in Molecular Biology Series, #174
Edition description:
2001
Pages:
438
Product dimensions:
10.00(w) x 7.00(h) x 1.00(d)

Table of Contents

Prefacev
Contributorsxi
IGenome and Transcript Analyses
1Epstein-Barr Virus: The B95-8 Strain Map3
2Analysis of Replication of oriP-Based Plasmids by Quantitative, Competitive PCR13
3Genetic Analysis and Gene Expression with Mini-Epstein-Barr Virus Plasmids23
4Construction of cDNA Libraries for the Analysis of the Structure of Complementary Strand Transcripts (CSTs)37
5Analysis of the Expression and Function of the EBV-Encoded Small RNAs, the EBERs, in Heterologous Cells45
6Visualizing EBV Expression Patterns by FISH67
IIViral Detection
7In Situ Detection of Epstein-Barr Virus DNA and Viral Gene Products79
8Phenotype Determination of Epstein-Barr Virus-Infected Cells in Tissue Sections93
9Detection of EBV Infection at the Single-Cell Level: Precise Quantitation of Virus-Infected Cells In Vivo103
10Detection and Discrimination of Latent and Replicative Herpesvirus Infection at the Single Cell Level In Vivo111
IIICulture Methods
11Virus Isolation119
12Generation of Lymphoblastoid Cell Lines (LCLs)125
13Cell Cycle Distribution of B-Lymphocytes and Cell Lines129
14Introduction of Plasmid Vectors into Cells Via Electroporation137
15Malignant Transformation and Immortalization Assays in Animal Cells Transfected with the BARF1 Gene147
16Transient Gene Expression and MACS Enrichment155
17In Vitro Assays to Study Epithelial Cell Growth165
18In Vitro Assays to Study Epithelial Cell Differentiation173
19Cell Sensitivity Assays: Quantitative Detection of Apoptotic Cells In Vitro Using the TUNEL Assay181
20Qualitative Detection of Apoptotic Cells Assessed by DNA Fragmentation189
IVImmune Assays
21Regression Assay199
22Generation of Polyclonal EBV-Specific CTL Cultures and Clones203
23Determination of Antigen and Fine Peptide Specificity of EBV-Specific CTLs209
24Limiting Dilution Assay213
VViral Protein Detection
25Antibodies for Detecting EBV Latent Proteins219
26Detection of EBV Latent Proteins by Western Blotting229
27Biosynthetic Radiolabeling of Virus Glycoproteins for Immunoprecipitation and Electrophoretic Analysis243
VIProtein-Protein, Protein-DNA, and Protein-RNA Interactions
28The Yeast Two-Hybrid Assay to Identify Interacting Proteins249
29Identification of Transactivation, Repression, and Protein-Protein Interaction Domains Using GAL4-Fusion Proteins259
30Magnetic DNA Affinity Purification of a Cellular Transcription Factor271
31Screening an Expression Library with a DNA Binding Site279
32Analysis of DNA Binding Proteins by Mobility Shift Assay289
33Analysis of RNA-Protein Interactions of the EBV-Encoded Small RNAs, the EBERs: In Vitro Assays297
VIIProtein Activity Assays
34Chimeric and Mutated Variants of LMP1: A Helpful Tool to Analyze the Structure-Function Relationship of a Pseudoreceptor313
35Assaying the Activity of Kinases Regulated by LMP1325
36In Vitro Assays for the Detection of Protein Tyrosine Phosphorylation and Protein Tyrosine Kinase Activities337
VIIITissue and In Vivo Protocols
37Analysis of Apoptosis in Tissue Sections347
38Considerations in Generating Transgenic Mice: DNA, RNA, and Protein Extractions from Tissues--Rapid and Effective Blotting361
39In Vivo Assay of Cellular Proliferation379
40Topical Chemical Carcinogen Treatment in Mice391
41Separation of Epidermal Tissue from Underlying Dermis and Primary Keratinocyte Culture401
42Selection and Enrichment of B Cells from Lymphoid Tissues411
43Detection of Immunoglobulin Gene Rearrangements423
Index433

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