Expression of HNF4α8 under the control of the P2 promoter. (A) Western blotting of whole cell extracts of the INS-1 P2/HNF4α8 cell lines (#1 and #2) with the HNF4α-antibody C-19 (Santa Cruz). Cells were cultured without (-) or with (+) 50 ng/ml tetracycline for 24 h. 20 μg of total protein were loaded per lane and the signal for the exogenous (exo) as well as endogenous (endo) HNF4α is marked. (B) Immunofluorescence of INS-1 P2/DD-HNF4α8#1 cells without (upper panel) or with (lower panel) induction with 50 ng/ml tetracycline and 1 μM Shield-1 for 45 h. HNF4α was detected using a myc-tag specific primary antibody and a Cy3-coupled secondary antibody (red). The cells were also stained with DAPI (blue) to visualize the total number of cells. The fusion protein is located in the nucleus comparable to the wild type HNF4α protein.