RESUMO
The ultrasensitive bio-barcode amplification assay (BCA) technique was developed for the specific detection of the outer-core protein VP7 of bluetongue virus (BTV). The target antigen VP7 was first captured by gold nanoparticles (GNPs) coated with polyclonal antibodies. Magnetic microparticles (MMP) coated with VP7 monoclonal antibody were then added to form a sandwich immuno-complex. After the immuno-complex was formed, signal DNA annealed to DNA strands covalently bound to the GNPs were released by heating and characterized by PCR and real-time fluorescence PCR. A detection limit of 0.1fg/ml was measured for purified VP7, seven orders of magnitude more sensitive than that of conventional antigen capture ELISA. The BCA demonstrated the same enhanced sensitivity for detecting BTV in serum samples from sheep. In the following work it is demonstrated that this assay is a highly sensitive method for the detection of BTV proteins that could be adapted to measure other proteins.
