1a.Objectives (from AD-416)
The long term goal of this project is the development of rapid, reliable, and sensitive diagnostic assays for the identification of zoonotic viruses that are suitable for use in point-of-care facilities, as well as in the field for use during times of natural or intentionally caused disease epidemics.
1b.Approach (from AD-416)
The feasibility of this goal will be demonstrated in the proposed research by the development of Au nanoparticle-mediated indirect and direct capture assays for arboviruses including Rift Valley Fever Virus (RVFV) or antibodies to these viruses. The detection technology is based upon surface enhanced Raman scattering (SERS) spectroscopy, an analytical tool that is well known for its high detection specificity and sensitivity and which affords several advantages over other nucleic acid detection and immunological technologies currently in use. This includes the reduction in assay costs by affording sensitivity without current amplification technologies and easy adaptation to point-of-care use. The primary focus of the proposed studies will be the demonstration and evaluation of the relative efficacies of alternative in vitro model assays, followed by the translation of these findings to the development of an optimized assay for detecting genetic material, antigen or antibodies to arboviruses.
Based on our past efforts in developing magnetic capture-based SERS assays for West Nile virus (WNV) nucleic acid detection, we have successfully demonstrated multiplex detection of WNV and Rift Valley Fever virus (RVFV) antigens. For these experiments, Au nanoparticles (GNPs) and paramagnetic nanoparticles (PMPs) coated with either polyclonal antibodies to the WNV envelope (E) protein or the RVFV N antigen were incubated with their respective target antigens, the GNP/antigen/PMP immuno-recognition complexes which developed were then retrieved from the solution reaction matrix by magnetic pull-down, and the concentrated target capture complex was interrogated using a 785 nm laser. Raman spectrum acquisitions specific for E antigen capture were provided by conjugating GNPs with the Raman reporter IR-792 and N antigen capture was provided by conjugating GNPs with the Raman reporter Nile Blue (NB). The lower limit of detection (LOD), for multiplex detection demonstrated that the SERS assay had significantly better sensitivities than characteristic for traditional ELISA’s.
This research supports NP103 Action Plan Components 1. Biodefense Research, and 3. Prevent and Control Zoonotic Diseases.
ADODR is directly involved in performance of the research and also monitors activities to evaluate research progress through site visits, meeting at conferences and through email and phone calls.