Rhodamine conjugated goat anti mouse

HAECs were plated on the multiwell plate containing a coverslip 24 hours prior to each treatment and oxLDL exposure. After appropriate treatment under oxLDL exposure, HAECs were washed with ice-cold PBS. Cells were fixed in 4% PFA for 20 minutes and permeabilized with 1% Triton X-100 in PBS. Prepared cells were blocked in 3% BSA in PBS for 1 hour and then incubated with each primary antibody overnight at 4°C as follows: anti-rabbit CD36 (1 : 100, Abcam), anti-mouse Trx (1 : 100, SCBT), and anti-rabbit LC3II (1 : 100, Cell Signaling Biotechnology). And then, each sample was incubated with fluorescence-conjugated secondary antibodies for 1 hour at room temperature. We used fluorescein isothiocyanate- (FITC-) conjugated donkey anti-rabbit (1 : 200, Jackson Immunoresearch, West Grove, PA, USA) or rhodamine-conjugated goat anti-mouse (1 : 200, Jackson Immunoresearch). Each sample was washed with PBS three times for 3 min each and then mounted with VECTASHIELD containing 4′,6-diamidino-2-phenylindole (DAPI) (Vector Laboratory, Carlsbad, CA, USA), and images were obtained using a Zeiss LSM 700 confocal microscope (Carl Zeiss, Thornwood, NY, USA). For staining the lysosome, LysoTracker (Molecular Probe, Eugene, OR, USA) was used using the manufacturer's instruction.

The cells used in this study were plated on the multi-well plate containing coverslip 24 h prior to each treatment and oxLDL exposure (50 µg/mL). After the appropriate treatment and oxLDL exposure, the cells were washed with ice-cold PBS. Cells were fixed in 4% PFA for 20 min and permeabilized with 1% TritonX-100 in PBS. The prepared cells were blocked in 3% BSA in PBS for 1 h and then incubated with each primary antibody overnight at 4 °C as follows: anti-rabbit p62 (1:100, Abcam), anti-mouse LAMP2 (1:100, SCBT), and anti-rabbit LC3II (1:100, Cell signaling biotechnology). Then, each sample was incubated with fluorescence-conjugated secondary antibodies for 1 h at room temperature. We used fluorescein isothiocyanate (FITC)-conjugated donkey anti-rabbit (1:200, Jackson Immunoresearch, West Grove, PA, USA) or rhodamine-conjugated goat anti-mouse (1:200, Jackson Immunoresearch). Each sample was washed with PBS three times for 3 min each and then mounted with Vectashield containing DAPI (Vector Laboratory, Carlsbad, CA, USA) and collected images using a Zeiss LSM 710 confocal microscope (Carl Zeiss, Thornwood, NY, USA).