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. 2016 Mar 17;8(4):689–697. doi: 10.1080/19420862.2016.1160988

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Figure 4. — Analysis of checkpoint complexes associated with the APC/C. (A) Schematic of the sequential purification of the APC/C complexes. HeLa cells arrested with nocodazole were harvested by mitotic shake-off and APC/C complexes purified with a monoclonal APC4 antibody and subsequently eluted by the antigenic peptide. This purified APC/C was subsequently used in BubR1 and pan-Mad2 purifications and analyzed by western blot. (B) Analysis of the APC/C complexes purified with the BubR1 antibody and the pan-Mad2 antibody. Representative of 3 independent experiments. (C) Quantification using Licor technology of the level of APC7, Mad2 and Cdc20 in the 2 purifications. Values were normalized to the level of BubR1 and the αBubR1 samples set to one. Average and standard deviation of 3 independent experiments indicated.