Cystic Fibrosis: Methods and Protocols consolidates cutting-edge in vitro, cellular, and whole animal laboratory protocols into an indispensable resource. From electrophysiology and cell biology, to animal models and gene therapy, this comprehensive set of methods provides the step-by-step instructions needed for investigators to incorporate new approaches into their research programs. Specific protocols describe new techniques for diagnosis, in vitro methods for the expression and functional analysis of CFTR, novel biochemical and cellular systems to determine how mutations subvert CFTR function, and in vivo protocols to examine how CFTR dysfunction produce multisystem pathology in both human and animal models.

Part I. Genetics of Cystic Fibrosis CFTR Mutation Detection by Multiplex Heteroduplex (mHET) Analysis on MDE Gel Julian Zielenski, Isabel Aznarez, Tuncer Onay, John Tzounzouris, Danuta Markiewicz, and Lap-Chee Tsui cDNA Microarrays for Pharmacogenomic Analysis of Cystic Fibrosis, Meera Srivastava Ofer Eidelman, and Harvey B. Pollard Natural Animal Models of Human Genetic Diseases Jeffrey J. Wine, Michael Dean, and Damjan Glavac Part II. CFTR Structure and Function: Structure, Gating, and Regulation Electrophysiological Approach to Studying CFTR Horst Fischer Quantitative Analysis of ATP-Dependent Gating of CFTR Allan Powe, Zhen Zhou, Tzyh-Chang Hwang, and Georg Nagel CFTR Regulation by Phosphorylation Tang Zhu, Deborah A. R. Hinkson, David Dahan, Alexandra Evagelidis, and John W. Hanrahan Transepithelial Measurements of Bicarbonate Secretion in Calu-3 Cells Robert J. Bridges Transepithelial Impedance Analysis of Chloride Secretion Ashvani K. Singh, Sangeeta Singh, Daniel C. Devor, Raymond A. Frizzell, Willy van Driessche, and Robert J. Bridges Studies of the Molecular Basis for Cystic Fibrosis Using Purified Reconstituted CFTR Protein Ilana Kogan, Mohabir Ramjeesingh, Canhui Li, and Christine E. Bear Probing CFTR Channel Structure and Function Using the Substituted-Cysteine-Accessibility Method Myles H. Akabas Methods for the Study of Intermolecular and Intramolecular Interactions Regulating CFTR Function Anjaparavanda P. Naren Fluorescent Indicator Methods to Assay Functional CFTR Expression in Cells Alan S. Verkman and Sujatha Jayaraman Expression, Folding, and Degradation Immunolocalization of CFTR in Intact Tissue and Cultured Cells Christopher R. Marino Analysis of CFTR Trafficking and Polarization Using Green Fluorescent Protein and Confocal Microscopy Bryan D. Moyer and Bruce A. Stanton CFTR Folding and Maturation in Cells Mohamed Benharouga, Manu Sharma, and Gergely L. Lukacs Isolation of CFTR: Chaperone Complexes by Co-Immunoprecipitation Geoffrey C. Meacham and Douglas M. Cyr CFTR Expression and ER-Associated Degradation in Yeast Yimao Zhang, Susan Michaelis, and Jeffrey L. Brodsky Manipulating the Folding Pathway of -F508 CFTR Using Chemical Chaperones Marybeth Howard and William J. Welch CFTR Degradation and Aggregation Michael J. Corboy, Philip J. Thomas, and W. Christian Wigley In Vitro Reconstitution of CFTR Biogenesis and Degradation Jon Oberdorf and William R. Skach In Vitro CFTR Folding Assays Rhesa D. Stidham, W. Christian Wigley, and Philip J. Thomas Analysis of CFTR Endocytosis by Cell Surface Biotinylation Kelly Weixel and Neil A. Bradbury Regulatory Complexes CFTR Regulation of ENaC Scott H. Donaldson, Elaine G. Poligone, and M. Jackson Stutts Yeast Two-Hybrid Identification and Analysis of Protein Interactions with CFTR Viswanathan Raghuram, Kenneth R. Hallows, and J. Kevin Foskett Biochemical Assays for Studying Indirect Interactions Between CFTR and the Cytoskeleton Peter J. Mohler, Patricia L. Kultgen, M. Jackson Stutts, and Sharon L. Milgram CFTR-Associated ATP Transport and Release Marie E. Egan Part III. Pathophysiology of Cystic Fibrosis Inflammatory Mediators in CF Patients, Jay B. Hilliard, Michael W. Konstan, and Pamela B. Davis Bacterial Colonization and Infection in the CF Lung Scott D. Sagel, Elaine B. Dowell, and Frank J. Accurso Antimicrobial Peptides and Proteins in the CF Airway, Alexander M. Cole and Tomas Ganz Bacterial-Epithelial Interactions Ruth Bryan and Alice Prince Thin-Film Measurements of Airway Surface Liquid Volume/Composition and Mucus Transport Rates In Vitro Robert Tarran and Richard C. Boucher Animal Models of Cystic Fibrosis Murine Models of CF Airway Infection and Inflammation James F. Chmiel, Michael W. Konstan, and Melvin Berger Analysis of Lipid Abnormalities in CF Mice Steven D. Freedman, Paola G. Blanco, Julie C. Shea, and Juan G. Alvarez Bioelectric Measurement of CFTR Function in Mice Barbara R. Grubb Xenograft Model of the CF Airway Mohammed Filali, Yulong Zhang, Teresa C. Ritchie, and John F. Engelhardt Development of Conditionally Immortalized Epithelial Cell Lines from CF and Non-CF Mice Calvin U. Cotton Technical Approaches to Analyze the In Vivo Ion Composition of Airway Surface Liquid Jean-Marie Zahm, Sonia Baconnais, Gérard Balossier, and Edith Puchelle Part IV. Novel Therapeutic Approaches for Cystic Fibrosis Design of Gene Therapy Clinical Trials in CF Patients Kimberly V. Curlee and Eric J. Sorscher Formulation of Synthetic Vectors for Cystic Fibrosis Gene Therapy John Marshall and Seng H. Cheng Adeno-Associated Viral Vectors for CF Gene Therapy Terence R. Flotte, Isabel Virella-Lowell, and Kye A. Chesnut