Skip to main content
. 2012 Mar 19;1(3):e4. doi: 10.1038/oncsis.2012.4

Figure 3.

Figure 3

Characterization of the NPM knockdown MDA-231 cells. (a) MDA-231 cells were transfected with NPM shRNA or control shRNA to generate stable NPM knockdown cells (MDA-231-NPM-KD) as described in materials and methods. Total cell lysates from MDA-231, -231-control shRNA and -231-NPM-KD cells were processed for western blot using NPM and actin antibodies. NPM levels were quantitated by normalizing the amount NPM in each cell line to the amount of actin in the same line relative to MDA-231 cells. The values for MDA-231-control shRNA and -231-NPM-KD transfectants are the average of multiple independent clones. The experiment was repeated at least three times and the displayed blot is from one typical experiment. (b) MDA-231-NPM-KD cells were processed for subcellular fractionation according to the Lamond's protocol and the cytoplasmic (C), nucleoplasmic (NP) and nucleolar (NU) fractions processed for immunoprecipitation with NPM antibodies followed by western blot with NPM and PG antibodies. Three independent clones were tested and the experiment was repeated at least three times. The data presented is for one typical experiment. (c) MDA-231, -231-NPM-KD and –231-NPM-KD+PG cells were grown to confluence, viewed under a phase contrast microscope and photographed using the × 20 objective. (d) Replicate cultures of MDA-231, -231-PG, -231-NPM-KD and -231-NPM-KD+PG cells were established at single cell density and cells were counted at 3, 5 and 7 days. Each time point represents the average of three independent experiments. The values for MDA-231-PG, -231-NPM-KD and -231-NPM-KD+PG transfectants are the average of at least three independent clones. The absence of error bars at some time points is because of the small differences among the experiments.