Das Xentry Bypass 19
if the system has already been saved license keys for xentry das, but for some reason require re-saving in some cases you cannot save the new keys without removing the old. deleting keys is performed by removing the files lic_key (daswis) and lic_key_2 (xentry) located along the way: c:documents and settingsall usersapplication data.
Das Xentry Bypass 19
xentry-kala and xentry-protamine (xp) were respectively diluted to 80 μmol/l in kcl-hepes and hepes-kcl buffer, and incubated at 37c for 5 min. after incubation, the solution was diluted to the desired concentration by dilution in pbs, followed by mixing with the appropriate transfection reagent at the desired ratio. the transfection reagent was prepared by mixing the appropriate volume of lipofectamine 2000 (invitrogen) with the appropriate amount of diluted xentry-kala or xentry-protamine (xp). the complexes were incubated for 5 min at room temperature before the mixture was added to cells and incubated for 48 h at 37c in a 5% co2 incubator.
the cells were harvested, washed in pbs and resuspended in the same buffer at a concentration of 2.0 104 cells/ml. the cells were then placed in a 96-well black-wall clear flat bottom plate (corning, new york, usa) with 100 μl cell suspension/well. the plates were centrifuged at 2000 rpm for 5 min and then incubated for 15 min at 37c in a co2 incubator (forma, thermo scientific, waltham, ma, usa). the supernatant was discarded and 100 μl of diluted cx43asodn was added to each well (100 μl/well) and incubated for 30 min at 37c in a co2 incubator. the supernatant was discarded and replaced with 100 μl of the diluted xentry-kala (xentry-protamine) (100 μl/well). the plate was then incubated for 48 h at 37c in a 5% co2 incubator. cells were fixed with 4% paraformaldehyde in pbs for 15 min and washed three times with pbs. the cell nuclei were stained with dapi at a concentration of 1.0 μmol/l for 10 min and the images were acquired using a fluorescence microscope (olympus, tokyo, japan).