The amino-terminal domain of the lamin B receptor is a nuclear envelope targeting signal

To remove the plasma membrane but keep the nuclear membranes intact, cells were permeabilized with digitonin as described by Adam et al. (1990) . A single petri dish of COS-7 cells was transfected with the desired plasmid, split into two dishes after 5 h and then grown as described above. After 24 h in culture, one dish was prepared for immunofluorescance microscopy as usual. The other dish of similarly transfected cells was washed three times with PBS and the cells were then permeabilized for 5 min with 40/xg/ml of digitonin (Calbiochem-Novabioehem Corp., La Jolla, CA) in PBS at 4 ~ (digitonin was diluted into PBS from a 20 mg/ml stock in DMSO). After permeabilization, cells were washed again with PBS and prepared for ira. munofluoreseence microscopy as usual except that the cells were not fixed and Triton X-100 was not present in solutions A and B. Immunofluorescenee microscopy was performed as above.