ER and Nucleus Staining of Aag2 Cells

Cells were plated on 18 mm × 18 mm coverslips in a 6-well cell culture plate 48 h previously coated with gelatin (0.2%, vol/vol) (62 (link), 86 (link)). To stain the ER, the cell culture medium was aspirated, and cells were washed with PBS once and incubated for 30 min at 26°C with 1 µM live ER-tracker red dye (Molecular Probes) diluted in the appropriate Aag2 cell culture medium. The ER-tracker solution was replaced by a 1/20,000 solution of SYTO-11 (Molecular Probes) DNA dye for 10 min at 26°C diluted in the appropriate Aag2 cell culture medium, washed twice with PBS, and cells were maintained in Aag2 culture medium during the confocal microscopy observations.