HeLa cells were cultured on cover slips and fixed with 4% paraformaldehyde in PBS (pH7.4) at room temperature for 10 min after exposed upon 10 uM choloroquine for 6 h. After washing three times with PBS, the cells were permeabilized with ice-cold methanol for 2.5 min. After three washes with PBS, the cells were incubated with blocking solution (5% BSA in PBS) for 1 h and then with primary antibody p62 overnight at 4ºC. The next day, the cells were washed five times for 5 min each time with PBS and then incubated with secondary antibody Goat Anti-Rabbit IgG H&L (Alexa Fluor 647) for 1 to 2 h. The cells were washed four times with PBS for 5 min each time, and DAPI stained for 15 min. After three washes with PBS, the coverslips were mounted on slides using Fluoro-GEL (Electron Microscopy Sciences). Images were taken using Keyence BZ-X700.
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Submitted Nov 01 2018
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