Cells (FANXA7−DOWN, PANXA7−UP, FANXA7−NC, PANXA7−NC, Hca-F and Hca-P) spread on poly-L-lysine-coated slides were fixed in 4% paraformaldehyde for 15 min. The cells were then blocked by incubation with goat serum (ZSGB-BIO, China) for one hour after incubation with rabbit anti-LEPR (Proteintech, USA, 1:200) and mouse anti-Annexin A7 (Abcam, USA, 1:200) overnight at 4 °C. The secondary antibody (DyLight 594 AffiniPure Donkey Anti-Rabbit/Mouse; Abbkine, USA, 1:50) was used at a 1:100 dilution for one hour at 37 °C. The cell nuclei were stained with DAPI (Beyotime, China) for 5 min at a concentration of 5 µg/ml and examined under a fluorescence microscope (Olympus, Japan).
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