(A) NFκB activation in resting and short-term activated CD4 T cells. Total CD4 T cells purified from BALB/c mouse spleen were used directly as resting cells or activated for 24hrs with anti-CD3/anti-CD28 antibodies before cotransfection with pRL3 encoding renilla luciferase along with plasmids pNK-Luc, pNFAT-luc, pCIS-K(neg. control) or pGL3-luc (positive control) by nucleofection (see methods), and cultured overnight. Dual luciferase activities were subsequently quantitated in cell lysates. Graph show normalized luciferase activity compared to the positive (pGL3-luc) control for resting and activated CD4 T cells, respectively. Results are compiled from three independent experiments. (B) NFκB activity in resting and activated memory CD4 T cells. Memory (CD44hi) CD4 T cells were fractionated from total CD4 T cells and transfected directly or after 24hrs activation with anti-CD3/anti-CD28 antibodies with pRL3 in combination with pGL3, pNFκB-luc or pCIS-K and dual luciferase activity quantitated and graphed as in (A). Results are representative of three independent experiments.