Progress 10/01/07 to 09/30/12
Outputs
Progress Report Objectives (from AD-416): To develop in vitro immunoassays to measure the efficacy of DNA vaccines for poultry. Approach (from AD-416): Develop in vitro cell culture system to measure gene expression, immune response, and vaccine efficacy. This project was initiated in 2007 as a result of a consortium grant provided by the Canadian Ministry of Agriculture and involves scientists from the Canadian Food Inspection Agency (CFIA), Ottawa laboratory (Fallowfield), CFIA�s Saskatoon laboratory, CFIA�s National Centre for Foreign Animal Disease laboratory in Winnipeg, the University of Saskatoon, and the Beltsville Agricultural Research Center, Beltsville, MD. In this final reporting period, the following research projects were conducted: 1) a series of murine hybridomas which secrete monoclonal antibodies against chicken interleukin (IL) 4 and interferon (IFN) gamma were developed; and 2) two monoclonal antibodies which showed the highest binding to the native chicken IL4 and IFN gamma were selected using an indirect enzyme-linked immunosorbant assay and provided to CFIA. These mouse monoclonal antibodies will be used to develop a new multiplex immunoassay to measure soluble cytokine production induced by infection with avian influenza virus. Furthermore, these two sets of mouse monoclonal antibodies will facilitate the delineation of the type of chicken T-helper cell responses, Th1 (cell-mediated) and Th2 (humoral).
Impacts
(N/A)
Publications
|
Progress 10/01/10 to 09/30/11
Outputs
Progress Report Objectives (from AD-416) To develop in vitro immunoassays to measure the efficacy of DNA vaccines for poultry. Approach (from AD-416) Develop in vitro cell culture system to measure gene expression, immune response, and vaccine efficacy. The project was initiated as a result of a consortium grant provided by the Canadian Ministry of Agriculture and involves scientists from CFIA�s Ottawa laboratory (Fallowfield), CFIA�s Saskatoon laboratory, CFIA�s National Centre for Foreign Animal Disease laboratory in Winnipeg, the University of Saskatoon, and the Beltsville Agricultural Research Center, USDA-ARS. In this reporting period, we developed an ELISA assay to evaluate the ability of recombinant H AI antigen to stimulate IFN-G. Mouse monoclonal antibodies were produced and IgG was purified to develop an antigen-capture assay to detect chicken IFN-G. This assay will be used to monitor embryo vaccination strategy against AI vaccine. This project is being monitored by monthly teleconferences and frequent meetings at the national and international meetings where both parties attend.
Impacts
(N/A)
Publications
|
Progress 10/01/09 to 09/30/10
Outputs
Progress Report Objectives (from AD-416) To develop in vitro immunoassays to measure the efficacy of DNA vaccines for poultry. Approach (from AD-416) Develop in vitro cell culture system to measure gene expression, immune response, and vaccine efficacy. This trust agreement was established in 2006 to develop an in ovo DNA vaccine strategy against avian influenza (AI). The project was initiated as a result of a consortium grant provided by the Canadian Ministry of Agriculture and involves scientists from Canadian Food Inspection Agency's (CFIA) Ottawa laboratory (Fallowfield), CFIA�s Saskatoon laboratory, CFIA�s National Centre for Foreign Animal Disease Laboratory in Winnipeg, the University of Saskatoon, and the Beltsville Agricultural Research Center, USDA-ARS. In this reporting period in vitro transfection system was used to express haemagglutinin (HA) gene in a mammalian vector. After validating HA antigen expression, fertilized eggs incubated for 18- day were injected in ovo with HA gene and on days 1, 2, and 3 days post- injection, the spleen, intestine, liver and lung were subjected to mRNA extraction and gene expression analysis. The results showed that the AI HA gene expression was significantly higher in the lungs of in ovo injected chicken groups compared with other groups. In the intestine, only a single inoculation group resulted in significantly higher levels of expression compared with control group. In liver and spleen samples, no significant changes were observed. These results indicated that with the injection of AI HA gene it is apparent that complex and dynamic responses of immune-related gene expression occurs in the different tissues/organs of different groups of chickens. This information will be used for development of embryo vaccination strategy against AI vaccine. Progress is monitored by written progress reports, regular teleconferences and meetings.
Impacts
(N/A)
Publications
|
Progress 10/01/08 to 09/30/09
Outputs
Progress Report Objectives (from AD-416) To develop in vitro immunoassays to measure the efficacy of DNA vaccines for poultry. Approach (from AD-416) Develop in vitro cell culture system to measure gene expression, immune response, and vaccine efficacy. Significant Activities that Support Special Target Populations This agreement with was established to develop an in ovo DNA vaccine strategy against avian influenza. The project was initiated as a result of a consortium grant provided by the Canadian Ministry of Agriculture and involves scientists from CFIA�s Ottawa laboratory (Fallowfield), CFIA�s Saskatoon laboratory, CFIA�s National Centre for Foreign Animal Disease laboratory in Winnipeg, the University of Saskatoon, and the Beltsville Agricultural Research Center, USDA-ARS. In this reporting period, in vitro transfection system was established to optimize the protein expression of H antigen in a mammalian vector. Following transfection of a chicken cell line with H5 gens, protein expression was evaluated at 24, 48 and 72 hrs using Western blot analysis. In three experiments, 24 hrs was the best time to detect H5 antigen. This information will be used for development of embryo vaccination strategy against AI vaccine.
Impacts
(N/A)
Publications
|
Progress 10/01/07 to 09/30/08
Outputs
Progress Report Objectives (from AD-416) To develop in vitro immunoassays to measure the efficacy of DNA vaccines for poultry. Approach (from AD-416) Develop in vitro cell culture system to measure gene expression, immune response, and vaccine efficacy. Significant Activities that Support Special Target Populations This research develops an in ovo DNA vaccine strategy against avian influenza. The Project was initiated as a result of a consortium grant provided by the Canadian Ministry of Agriculture and involves scientists from USDA-ARS, CFIA�s Ottawa laboratory (Fallowfield), CFIA�s Saskatoon laboratory, CFIA�s National Centre for Foreign Animal Disease laboratory in Winnipeg, and the University of Saskatoon. ARS scientists provide expertise in the area of embryo vaccination technology and molecular immunology to carry out a joint research project entitled "Development of an in vitro test for the evaluation of the potency of an avian influenza DNA vaccine for chickens." During the first year, real-time PCR for various chicken cytokines and chemokines were optimized and DNA vectored recombinant vaccines were constructed. This project is being monitored by monthly teleconferences and frequent meetings at the national and international meetings where both parties attend.
Impacts
(N/A)
Publications
|
Progress 10/01/06 to 09/30/07
Outputs
Progress Report Objectives (from AD-416) To develop in vitro immunoassays to measure the efficacy of DNA vaccines for poultry. Approach (from AD-416) Develop in vitro cell culture system to measure gene expression, immune response, and vaccine efficacy. Significant Activities that Support Special Target Populations This report documents research conducted under a Trust Fund Agreement between ARS and the Canadian Food Inspection Agency (CFIA). Additional details of the research can be found in the associated in-house project 1265-31320-072-00D, �Development of Functional Genomic and Immunologic Approaches to Control Avian Mucosal Pathogens�. This is a trust agreement that is being established to evaluate novel DNA vaccination strategy. The planned research will start as soon as the trust agreement is finalized. Monitoring progress on this cooperative agreement will be in emails, phone calls, and periodic conversations at scientific meetings.
Impacts
(N/A)
Publications
|
|