Transfection of Ramos cells with inducible NF-κB modulators. (A) Immunoblot analysis of transfected Ramos cells untreated or treated with doxycycline (0.5 μg/m) for 48 hours. Cell extracts were analyzed for CA-IKK2, for TD-IκBα, and as a control for actin expression. (B) EMSA of cell extracts shows induction of NF-κB activity after doxycycline addition. Oct1 DNA was used as control for the integrity of the extracts. (C) Morphologic examination of transfected Ramos. Bright field and fluorescence microscopy of cells treated with doxycycline for 5 days. Images were taken with a Leica DMIRBE microscope 10× NA = 0.3 PH1 acquired with a Hamamatsu digital camera C4742-95 (Hamamatsu Photonics) and Open Lab software Version 4.0.4 (Improvision). (D) Proliferation assay of Ramos expressing pRTS-GFP (◇), pRTS-TD-IκBα (□), or pRTS-CA-IKK2 (▲). GFP-positive cells were counted by flow cytometer at the indicated time points after doxycycline addition. Error bars represent SDs from triplicate experiments.