Poxviruses are large DNA viruses that replicate in discrete locations in the cytoplasm of infected cells called viral factories. Because the host cell DNA replication machinery is located in the nucleus, poxviruses encode many of the
proteins required for their own DNA replication, including
a DNA polymerase. Although many if not all of the
enzymes that are required for
viral DNA replication have been identified, the actual mechanism of poxvirus DNA replication remains unclear. Two
monoclonal antibodies and a polyclonal antibody against
vaccinia virus DNA polymerase were produced and characterized for use as tools to investigate the mechanism of virus DNA replication. Although the
monoclonal antibodies were not suitable for Western blotting, the polyclonal antibody was able to detect the
protein in infected cell lysates using this method. In contrast, while the polyclonal antibody did not recognize the
DNA polymerase when used for immunofluorescence microscopy, the
monoclonal antibodies were able to detect the polymerase in
vaccinia viral factories. In addition, one of these
antibodies also stained viral factories produced by
cowpox and
ectromelia, two closely related viruses. Finally, all three
antibodies were able to immunoprecipitate
vaccinia DNA polymerase from infected cell lysates. These
antibodies will be useful in experiments designed to describe more fully the role of the
viral DNA polymerase in DNA replication of vaccinia virus.