Retinal Gene Therapy

1. Front Matter

   Pages i-xvi

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2. Retinal Gene Therapy Vector Production and Biological Assays In Vitro

   1. Front Matter

      Pages 1-1

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   2. Small Scale Production of Recombinant Adeno-Associated Viral Vectors for Gene Delivery to the Nervous System

      • Joost Verhaagen, Barbara Hobo, Erich M. E. Ehlert, Ruben Eggers, Joanna A. Korecka, Stefan A. Hoyng et al.

      Pages 3-17

   3. Small and Micro-Scale Recombinant Adeno-Associated Virus Production and Purification for Ocular Gene Therapy Applications

      • Christopher A. Reid, Daniel M. Lipinski

      Pages 19-31

   4. Design and Development of AAV-based Gene Supplementation Therapies for Achromatopsia and Retinitis Pigmentosa

      • Christian Schön, Elvir Becirovic, Martin Biel, Stylianos Michalakis

      Pages 33-46

   5. Development of Multigenic Lentiviral Vectors for Cell-Specific Expression of Antiangiogenic miRNAs and Protein Factors

      • Anne Louise Askou, Thomas J. Corydon

      Pages 47-60

   6. Design and In Vitro Use of Antisense Oligonucleotides to Correct Pre-mRNA Splicing Defects in Inherited Retinal Dystrophies

      • Alejandro Garanto, Rob W. J. Collin

      Pages 61-78

   7. Three-Dimensional Co-Culture Bioassay for Screening of Retinal Gene Delivery Systems

      • Ding Wen Chen, Kathleen Pauloff, Marianna Foldvari

      Pages 79-88

   8. Retinal Gene Therapy for Choroideremia: In Vitro Testing for Gene Augmentation Using an Adeno-Associated Viral (AAV) Vector

      • Maria I. Patrício, Robert E. MacLaren

      Pages 89-97

3. Assays for Gene Augmentation and Editing In Vivo on Rodent and Macaque Retina

   1. Front Matter

      Pages 99-99

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   2. In Vivo Electroporation of Developing Mouse Retina

      • Jimmy de Melo, Seth Blackshaw

      Pages 101-111

   3. Methods for In Vivo CRISPR/Cas Editing of the Adult Murine Retina

      • Sandy S. Hung, Fan Li, Jiang-Hui Wang, Anna E. King, Bang V. Bui, Guei-Sheung Liu et al.

      Pages 113-133

   4. AAV Gene Augmentation Therapy for CRB1-Associated Retinitis Pigmentosa

      • C. Henrique Alves, Jan Wijnholds

      Pages 135-151

   5. Dual AAV Vectors for Stargardt Disease

      • Ivana Trapani

      Pages 153-175

   6. Optogenetic Retinal Gene Therapy with the Light Gated GPCR Vertebrate Rhodopsin

      • Benjamin M. Gaub, Michael H. Berry, Meike Visel, Amy Holt, Ehud Y. Isacoff, John G. Flannery

      Pages 177-189

   7. CRISPR Repair Reveals Causative Mutation in a Preclinical Model of Retinitis Pigmentosa: A Brief Methodology

      • Wen-Hsuan Wu, Yi-Ting Tsai, Sally Justus, Galaxy Y. Cho, Jesse D. Sengillo, Yu Xu et al.

      Pages 191-205

   8. In-Depth Functional Analysis of Rodents by Full-Field Electroretinography

      • Vithiyanjali Sothilingam, Regine Mühlfriedel, Naoyuki Tanimoto, Mathias W. Seeliger

      Pages 207-213

   9. Advanced Ocular Injection Techniques for Therapy Approaches

      • Regine Mühlfriedel, Marina Garcia Garrido, Christine Wallrapp, Mathias W. Seeliger

      Pages 215-223

   10. Neutralizing Antibodies Against Adeno-Associated Virus (AAV): Measurement and Influence on Retinal Gene Delivery

       • Mélissa Desrosiers, Deniz Dalkara

       Pages 225-238

   11. Screening for Neutralizing Antibodies Against Natural and Engineered AAV Capsids in Nonhuman Primate Retinas

       • Timothy P. Day, Leah C. Byrne, John G. Flannery, David V. Schaffer

       Pages 239-249

This detailed volume describes a spectrum of methods and protocols that can be used for the bench-to-bedside development and evaluation of retinal gene therapy. Methods for the successful delivery of these gene therapy vector systems to the retina are examined, as well as assays to test the efficacy in vitro in cell cultures, for gene augmentation and gene editing in vivo on rodents, pigs, and monkey retinas, and on human retinal explants as well as in human clinical studies. Written for the highly successful Methods in Molecular Biology series, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible protocols, and tips on troubleshooting and avoiding known pitfalls. 

Authoritative and practical, Retinal Gene Therapy: Methods and Protocols provides a wide range of readers from students to research experts with vital information on ocular gene therapy vector technology, in vitro and in vivo biological assays, and clinical protocols, to promote further studies for the benefit of children and adults with inherited retinal disease.

• Gene therapy vector
• Retinal cell types
• Gene augmentation
• Cell cultures
• Gene silencing
• Vector production
• AAVs

• Department of Ophthalmology, Leiden University Medical Center (LUMC), Leiden, The Netherlands

  Camiel J.F. Boon, Jan Wijnholds

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