ALDEFLUOR assay

CSCs were also identified by ALDEFLUOR assay (StemCell Technologies) by measuring the activity of aldehyde dehydrogenase (ALDH). MCF-7 cells were treated with 10 μM of protein kinase inhibitors for 24 h, and the proportion of ALDH-positive cells was analyzed with ALDEFLUOR assay. Briefly, MCF-7 cells were suspended with buffer containing BODIPY-aminoacetaldehyde and incubated for 45 min at 37 °C. For each sample, cells were incubated with or without diethylaminobenzaldehyde (DEAB), an ALDH inhibitor. ALDH-positive and negative cells were sorted using the FACSCalibur flow cytometer (BD Bioscience).