Transfection of anti-rabies antibodies in N2A cell line (murine neuroblastoma) and intracellular neutralization for use as antiviral

• Grant number: 11/06585-5
• Support Opportunities: Regular Research Grants
• Start date: July 01, 2011
• End date: June 30, 2013
• Field of knowledge: Agronomical Sciences - Veterinary Medicine - Preventive Veterinary Medicine
• Principal Investigator: Juliana Galera Castilho Kawai
• Grantee: Juliana Galera Castilho Kawai
• Host Institution: Faculdade de Medicina Veterinária e Zootecnia (FMVZ). Universidade de São Paulo (USP). São Paulo , SP, Brazil
• Associated researchers: Paulo Eduardo Brandão

Abstract

Rabies is an acute, fatal zoonosis that affects mammals. It represents a serious public health problem, particularly in developing countries where canine rabies is endemic, and is responsible for around 55,000 deaths every year. The search for drugs and alternative mechanisms against rabies virus is therefore of worldwide interest. The objective of this study is to investigate whether anti-rabies antibodies can be transfected using a cationic reagent (lipofectamine) in an in vitro system and to examine the effectiveness of intracellular neutralization by anti-rabies antibodies as an antiviral therapy. For this end, different variants of the rabies virus adapted to N2A (murine neuroblastoma) cells and molecularly characterized by DNA sequencing will be used. Transfection of anti-rabies antibodies using lipofectamine will be carried out with anti-rabies nucleocapsid conjugate, and the success of the transfection will be evaluated by fluorescence microscopy. Once the suitability of lipofectamine for transfection of anti-rabies antibodies has been demonstrated and the isolates titrated, intracellular neutralization of the rabies virus will be tested using anti-rabies nucleocapsid polyclonal antibody. Studies using antibody transfection and intracellular neutralization of the rabies virus in cell culture as the basis of potential antiviral therapies will enable improvements to be made to this technique by providing a greater understanding of its efficiency and the way in which it operates. This in turn will provide the basis for studies of the efficiency of this technique in animal models, an essential step if safer, more effective protocols for treating rabies are to be developed and the chances of survival of rabies patients are to be increased. (AU)