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Protocols and Applications

Protocol

Aspects of Gene Therapy Products Using Current Genome-Editing Technology in Japan

The development of genome-editing technology could lead to breakthrough gene therapy. Genome editing has made it possible to easily knock out or modify a target gene, while current gene therapy using a virus vector or plasmid hampering modification with respect to gene replacement therapies. Clinica... More


Protocol

Advances in Gene Therapy for Diseases of the Eye

Over the last few years, huge progress has been made with regard to the understanding of molecular mechanisms underlying the pathogenesis of neurodegenerative diseases of the eye. Such knowledge has led to the development of gene therapy approaches to treat these devastating disorders. Challenges re... More


Protocol

Gene Therapy for Hemoglobinopathies

Gene therapy for β-thalassemia and sickle-cell disease is based on transplantation of genetically corrected, autologous hematopoietic stem cells. Preclinical and clinical studies have shown the safety and efficacy of this therapeutic approach, currently based on lentiviral vectors to transfer a β-gl... More


Protocol

Long Term Storage, Recycling and Simplified Calcium Phosphate Transfection Protocols for Cell Cultures

Long term storage of cells is vital for use in the future experiments. Current methods of storing cells under liquid nitrogen vapor is costly, and requires regular monitoring and filling of the storage tank. We stored several cell lines in -80oC freezer over a decade and revived them successfully. S... More


Protocol

Zebrafish Tumor Graft Transplantation to Grow Tumors In Vivo That Engraft Poorly as Single Cell Suspensions

Angiosarcoma is a clinically aggressive tumor with a high rate of mortality. It can arise in vascular or lymphatic tissues, involve any part of the body, and aggressively spread locally or metastasize. Angiosarcomas spontaneously develop in the tp53 deleted (tp53del/del) zebrafish mutant. However, e... More


Protocol

An Automated Low-Cost Swim Tunnel for Measuring Swimming Performance in Fish

The study of swimming behavior is an important part of fish biology research and the swim tunnel is used to study swimming performance as well as metabolism of fish. In this investigation, we have developed a user-friendly, automated, modular, and low-cost swim tunnel that permits to study the perfo... More


Protocol

A Rabbit Femoral Condyle Defect Model for Assessment of Osteochondral Tissue Regeneration

Osteochondral tissue repair represents a common clinical need, with multiple approaches in tissue engineering and regenerative medicine being investigated for the repair of defects of articular cartilage and subchondral bone. A full thickness rabbit femoral condyle defect is a clinically relevant mo... More


Protocol

Cartilage Assessment Requires a Surface Characterization Protocol: Roughness, Friction, and Function

The surface of articular cartilage is integral to smooth, low-friction joint articulation. However, the majority of cartilage literature rarely includes measurements of surface characteristics and function. This may, in part, be due to a shortage of or unfamiliarity with fast, nondestructive, and, p... More


Protocol

Optimized Protocol for Accurate Titration of Adeno-Associated Virus Vectors

Adeno-associated virus (AAV) is currently the most popular gene delivery vector for in vivo gene therapy. However, variability in titration methods between different laboratories affects the reproducibility of experiments and evaluation of safety and efficacy in clinical trials. We describe an optim... More


Protocol

Single-Use Capture Purification of Adeno-Associated Viral Gene Transfer Vectors by Membrane-Based Steric Exclusion Chromatography

We present membrane-based steric exclusion chromatography (SXC) as a universal capture step for purification of adeno-associated virus (AAV) gene transfer vectors independent of their serotype and surface characteristics. SXC is performed by mixing an unpurified cell culture supernatant containing A... More