2012 Annual Report
1a.Objectives (from AD-416):
The goal of this collaborative research project is to produce camel monoclonal antibodies (cMAb) against Foot-and-Mouth Disease Virus (FMDV) non-structural proteins (NSP) and virus particles (SP). USDA, ARS and Ben Gurion University scientists will design a reagent for viruses that will target three relevant FMDV serotyes including O, A and the South African Territories (SAT) types of Foot-and-Mouth Disease Virus (FMDV). An additional objective includes an attempt to assemble diagnostics assays using selected virus-specific cMAb developed in this project. Currently, there are no effective “in house” and affordable surveillance diagnostic test widely available in sub-Saharan Africa.
1b.Approach (from AD-416):
Virus specific camel monoclonal antibodies (cMAbs) in camelids directed against the Foot-and-Mouth Disease Virus (FMDV) structural proteins (SP) and non-structural proteins (NSPs) will be produced. Using an eukaryotic expression system tailored to the FMDV proteins, reactive hybridoma clones to the capsid, the 3D and 3ABC proteins will be screened and selected. Additionally, efforts will be aimed at the development of suitable and easy to deploy DIVA (differentiation between infected and vaccinated animals) (DIVA) tests utilizing the camel MAbs in a test platform. The formulated test/s will be bench standardized to assess the rapid and accurate detection of anti-NSP and FMDV detection. The specificity, stability and affinity characteristics of camel MAbs make them excellent choices in biosensor applications and they will potentially be more stable reagents when applied in a validated test.
The goal of the program developed at Ben Gurion University with ARS, PIADC is to generate camel monoclonal antibodies for production of thermostable kits for diagnosis of Foot-and-Mout Disease (FMD) in Central Africa.
In FY 2012 we have accomplished the following; first, we purchased two camels for the purpose of inoculating them with FMD antigens. We also provided for food, care and handling of the camels for the first year. These costs were a significant portion of the seed funds received from ARS, PIADC. The camels were inoculated with inactivated crude viral preps provided by the ARS, PIADC along with a marker protein that we used to monitor the immune response. The immune response of the camels was weak by ELISA to both marker and immunogen. Ultracentrifugation gradient purification failed to yield sufficiently pure material for further inoculations. As such, we decided to use the remainder of the seed funding to begin expressing recombinant proteins of FMD in bacteria for inoculation of the camels. Clones of the VP1 gene for FMD strains A24, O, SAT 1 and South African Territories (SAT) 2 were synthesized, sequenced and expressed. We managed to get a small amount of soluble form of protein and worked that up for further inoculations. Most of the expressed protein for these clones is insoluble and we purified this material and performed refolding on it for future inoculation.
Activities slated for FY 2013 are on hold right now due to lack of funds for this work. As such, the goals of this project have not yet been met according to the initial plan due to the lack of funds. The original goals as stated in the agreement consisted of steps to harvest camel lymphocytes from inoculated animals for the purpose of producing anti-FMD specific camel monoclonal antibodies for thermostable kits for FMD diagnostics in Africa. We have successfully initiated this project with the seed funding and have produced several FMDV recombinant proteins. Once further funding is forthcoming the camels will be inoculated with these recombinant proteins along with others to be produced. We will then isolate antibodies and produce kits as initially planned.
No technologies were transferred or publications prepared during FY 2012.