Over the last decade, frequent epidemic outbreaks of
hand, foot and mouth disease have been observed in the Asia-Pacific region.
Hand, foot and mouth disease is caused by different viruses from the enterovirus family, mainly coxsackievirus A16 and enterovirus 71 (EV71) from the human enterovirus A family. Severe disease and neurological complications are associated more often with EV71
infection, and can lead occasionally to fatal brain stem
encephalitis in young children. The rapid progression and high mortality of severe
hand, foot and mouth disease makes the direct detection of
antigens early in
infection essential. The best method for virus detection is the use of specific
monoclonal antibodies. The generation and characterization of a
monoclonal antibody specific for the 3D polymerase of human enterovirus A and the development of a virus detection dot blot assay are described. A recombinant 3CD
protein from EV71 C4 strain was used as an immunogen to generate
monoclonal antibodies (MAbs). Screening of hybridoma cells led to the isolation of
monoclonal antibody 4B12 of the
immunoglobulin IgG1 isotype. MAb 4B12 recognizes the linear
epitope DFEQALFS close to the active site of the 3D polymerase, corresponding to
amino acid positions 53-60 of 3D and 1784-1791 of enterovirus 71
polyprotein. The presence of 3D polymerase and its precursor 3CD
proteinase in purified virus particles was confirmed. MAb 4B12 was used successfully to detect all enterovirus 71 subgenotypes in a denaturing dot blot assay with a sensitivity of 10 pg of 3D
protein and 10(4) tissue culture infective dose of virus particles.