Constitutive activation of NF-κB in ovarian cancer cell lines.

<p>(A) Five micrograms of nuclear extracts from the indicated cell lines were subjected to EMSA using ³²P-labeled oligonucleotides containing an NF-κB binding sequence or Oct-1-binding sequence as probes. (B) For competition assay nuclear samples were preincubated with 100-fold molar excess cold oligonucleotide before adding the labeled probe (left panel). DNA-binding NF-κB components in the indicated cell lines were determined by super-shift EMSA (right panel). Nuclear extracts (5 µg) from the indicated cell lines were preincubated for 30 minutes with purified mouse IgG, anti-p50 or anti-c-Rel antibodies, pre-immune (PI), anti-p52, anti-RelA or anti-RelB sera, and then subjected to EMSA with the NF-κB-specific probe. (C and D) Nuclear (10 µg), cytoplasmic (30 µg) or whole-cell (30 µg) extracts were subjected to SDS-PAGE followed by immunoblottings with the indicated antibodies.