HIV Protocols

by Prasad, Vinayaka R., Ph.D.; Kalpana, Ganjam V.; Walker, John M.

• ISBN13: 9781588298591
• ISBN10: 1588298590
• Edition: 2nd
• Format: Hardcover
• Copyright: 2008-12-05
• Publisher: Humana Pr Inc

Summary

Despite major advances in HIV treatment, many areas require more study, in order to create efficacious, potent antiretrovirals that can suppress viral load completely and durably without toxic side effects, to define unknown drug targets and fine-tune known targets, and to better understand the interplay between viral and host factors. In "HIV Protocols, Second Edition", expert researchers provide clear, state-of-the-art methods for the study of HIV. Directed toward three specific goals, this text aims to document up-to-date protocols for select aspects of HIV biology, to bring together both virological and immunological approaches in a single, convenient volume, and to present a comprehensive account of a range of techniques not available in any existing HIV protocol book. As a volume in the highly successful Methods in Molecular Biology'„¢ series, the chapters include brief introductions to the subject, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and Notes sections containing priceless tips on troubleshooting and avoiding known pitfalls.Comprehensive and cutting-edge, "HIV Protocols, Second Edition" is an ideal guide to the wide array of techniques used in fundamental or applied research into the biology and pathogenesis of HIV-1.

Table of Contents

Preparation of Virus Particles and Their Analysis

Methods for Viral RNA Isolation and PCR Amplification for Sequencing of Near Full-Length HIV-1

Purification of HIV-1 Virions by Subtilisin Digestion or CD45 Immunoaffinity Depletion for Biochemical Studies

Calculating HIV-1 Infectious Titer Using a Virtual TCID50 Method

Methods to Study HIV-1 Replication

Early Events

Cell-free Assays for HIV-1 UncoatingChristopher Aiken

Real-Time PCR Analysis of HIV-1 Replication Post Entry Events

Analysis of 2-LTR Circle Junctions of Viral DNA in Infected Cells

HIV-1 Recombination: An Experimental Assay and a Phylogenetic Approach

Methods of Preparation and Analysis of Intracellular Reverse Transcription Complexes

Analysis of Viral and Cellular Proteins in HIV-1 Reverse Transcription Complexes by Co-immunoprecipitation

Isolation and Analysis of HIV-1 Preintegration Complexes

Bisarsenical Labeling of HIV-1 for Real-Time Fluorescence Microscopy

Late Events

Methods for the Study of HIV-1 Assembly

Assembly of Immature HIV-1 Capsids using a Cell-Free System

Preparation of Recombinant HIV-1 Gag Protein and Assembly of Virus-like Particles in vitro

Methods for the Analysis of HIV-1 Nucleocapsid Protein Interactions with Oligonucleotides

Methods for Analysis of Incorporation and Annealing of tRNALys in HIV-1

Specialized Approaches to Study HIV-1 Biology and Pathogenesis

Somatic Cell Genetic Analysis to Identify HIV-1 Host Restriction Factors

Rapid, Controlled and Intensive Lentiviral Vector-based RNAi

Reverse Two Hybrid Screening to Analyze Protein-protein Interaction of HIV-1 and Cellular Proteins

Methods to Study Monocyte migration induced by HIV-infected cells

Novel Mouse Models for Understanding HIV-1 Pathogenesis

Immunological Studies of HIV

Mucosal Immunology

Mucosal Antibody Responses to HIV

Isolating Mucosal Lymphocytes from Biopsy Tissue for Cellular Immunology Assays

Measuring T cell Responses via Flow Cytometry

Quantifying HIV-1-specific CD8+ T-cell Responses Using ELISPOT and Cytokine Flow Cytometry

Multiparameter Flow Cytometry Monitoring of T cell Responses

Antiviral Responses

Measuring HIV Neutralization in a Luciferase Reporter Gene Assay

Assessing the Antiviral Activity of HIV-1-specific cytotoxic T lymphocytes

Methods for Quantitating Antigen-specific T Cell Responses Using Functional Assays in Rhesus Macaques

Isolation of Drug-Resistant Mutant HIV Variants Using Tissue Culture Drug Selection

Virus Evolution as a Tool to Study HIV-1 Biology

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