Visualizing LDLR Localization in HepG2 Cells

HepG2 cells were cultured in glass-bottom cell culture plates at 37 °C for 12 h. After 6 h of starvation with a serum-free medium, cells were overlaid with medium containing R and Q for another 24 h. Following this, the cells were washed with PBS, fixed with paraformaldehyde (4%) for 15 min at room temperature, and permeabilized with Triton X-100 (0.1%) for 5 min, and then cells were blocked in BSA (5% in PBS) for at least 1 h and incubated with antibody against LDLR (1:250 in BSA) overnight at 4 °C. After being washed with PBS three times, cells were immunostained with secondary fluorescence antibody for 1 h at room temperature [52 (link)]. The cell nucleus was visualized with DAPI. After washing with PBS three times, cells were examined using a confocal microscopy, and the fluorescence intensity was analyzed with Fluoview viewer software (Olympus) [47 (link)].