4 6 diamidino 2 phenylindole dapi staining solution

MN (purity > 98%) and HK (purity > 98%) were purchased from ChemFaces (Wuhan, PRC). Ko143 (purity 96%) was obtained from Enzo Life Sciences, Inc. (Farmingdale, NY, USA). MXR (purity 99%) was obtained from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA, USA). Dimethyl sulfoxide (DMSO), formic acid, sodium dodecyl sulfate (SDS), 3-(4′,5′-dimethylthiazol-2′-yl)-2,5-diphenyltetrazolium bromide (MTT), butylparaben and triton X-100 were supplied by Sigma-Aldrich Chemical Co. (St. Louis, MO, USA). Fetal bovine serum (FBS) was obtained from Biological Industries Inc. (Kibbutz, Beit Haemek, Israel). Penicillin-Streptomycin-Glutamine, Dulbecco’s modified Eagle medium (DMEM), 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid, Hank’s buffered salt solution (HBSS)and trypsin/EDTA were purchased from Invitrogen (Grand Island, NY, USA). Antibodies against ABCG2 and GAPDH were purchased from GeneTex (San Antonio, TX, USA). Polyvinylidene fluoride transfer membranes (Immobilon P) and chemiluminescence (ECL) were obtained from Millipore Corp. (Bedford, MA, USA). 4′,6-diamidino-2-phenylindole (DAPI) staining solution and goat anti-rabbit IgG H&L (DyLight™ 488) antibody were purchased from Abcam (Cambridge, UK). Acetonitrile (ACN) and methanol with LC grade were obtained from Mallinckrodt Baker (Phillipsburg, NJ, USA). Milli-Q plus water (Bedford, MA, USA) was used throughout this study.

A BrdU cell proliferation detection kit (Abcam, Shanghai, China) was used to detect the RB cell proliferation. RB cells in the logarithmic growth phase were prepared into a single-cell suspension and seeded in 24-well plates (1 × 10⁵ cells/well). The BrdU labeling reagent was added according to the manufacturer's instructions, and the cells were incubated for 24h. After that, the cells were fixed and incubated with an anti-BrdU antibody (Abcam, Shanghai, China). Next, the cells were stained with 4′, 6-Diamidino-2-Phenylindole (DAPI) staining solution (Abcam, Shanghai, China). The number of BrdU-positive cells and the total number of DAPI-positive cells in three fields were randomly selected and counted under a fluorescence microscope (Olympus, Tokyo, Japan). Cell proliferation rate = number of BrdU-positive cells/DAPI-positive cells