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Fig. 5. Topo II activity in knockdown cells. The enzymatic activity of topo II was analyzed by a decatenation assay using kinetoplast DNA as a substrate. We inoculated 4x105 cells and prepared extracts for topo II assays from control (A), topo II{alpha}-knockdown (B) and topo II{alpha}ß-knockdown (C) cells 3 days after transfection. In each reaction mixture, a serial twofold dilution of topo II extracts was added. Lane 1: 1/1280 dilution; lane 2: 1/640; lane 3: 1/320; lane 4: 1/160; lane 5: 1/80; lane 6: 1/40; lane 7: 1/20; lane 8: 1/10; lane 9: 1/5; lane 10: undiluted extract. C1: no topo II extract; C2: undiluted extract without ATP; C3: 1/5 dilution without ATP; M1: linear kinetoplast DNA marker (**); M2: decatenated kinetoplast DNA marker (*). (D) Crude extracts immunoblotted using the anti-topo II{alpha}ß antibody 6H8.