Monoclonal antibody to RNA polymerase I of the silkworm

RNA polymerase I has been purified from the silk glands of the silkworm and antibodies obtained from immunized BALB/C mice. Hybridomas were obtained by fusing spleen cells from the same mice with SP-2/0 cells. One of the cloned hybridomas was injected into mice to produce ascitic fluid. Both polyclonal and monoclonal antibodies were characterized for their reactivity with polymerase I by double immunodiffusion, enzyme-linked immunosorbent assay, and enzyme inhibition. The monoclonal antibody is active even at 1:100,000 dilution, is specific for the second largest subunit of RNA polymerase I, and does show some cross-reactivity with polymerases II and III. The monoclonal antibody when coupled to Sepharose binds polymerase I from crude extracts, and the bound polymerase I can be eluted. This antibody has been employed to localize RNA polymerase to the nuclei of Chironomus salivary glands. This and other monoclonal antibodies will be of considerable help as probes for the study of structure and function of RNA polymerases during active transcription.