Microplate autoreader

786‐O or A498 cells were seeded in 96‐well plates at a concentration of 2 × 10³/well. CCK8 solution (20 μL; Dojindo) was added to each well and mixed for 2 hours. Microplate Autoreader (ThermoFisher) was used to measure absorbance at 450 nm every 24 hours. For colony formation experiments, 786‐O or A498 cells were seeded on six‐well plates at a concentration of 1 × 10³/well with RPMI 1640 medium. Fourteen days later, the colonies were fixed in methanol and stained with crystal violet (0.1%). Visible colonies were counted and photographed under a light microscope (Bx41, Olympus).
5‐Ethynyl‐2′‐deoxyuridine (EdU) cell proliferation assay kit (RiboBio) was used to detect cell proliferation of 786‐O or A498 cells. Cultured 786‐O or A498 cells (200 μL of 2 × 10⁴/mL) were incubated with 50 µmol/L EdU for 8 hours. After fixation with 70% alcohol and permeabilization with Triton X‐100, the cells were then incubated with Apollo Staining reaction liquid (Click‐iT™ Edu Apollo Stain Kit (Invitrogen) to label the cells. Nuclei were stained with DAPI. Immunostaining was visualized and photographed under a fluorescent microscope (Olympus inverted microscope IX71).

HCT116, HT29, HCT116/R, and HT29/R (5X10³ cells/well) were seeded in 96-well plates, and treated with 1, 2, 4, 8, 16, 32, 64, or 128 μM oxaliplatin for 24 hours. Cells were then incubated with CCK8 (Cell Counting Kit-8) (Dojindo, Tokyo, Japan) for another 2 hours, and the absorbance at 450 nm was measured using Microplate Autoreader (Thermo Fisher) according to previous study [15 (link)]. Moreover, HCT116/R and HT29/R cells were seeded in 96-well plates, and then transfected with shNC, shWDR62-1# or shWDR62-2# (GenePharma, Suzhou, China) using Lipofectamine 2000 (Gibco, Carlsbad, CA, USA) for 48 hours. Cells were then also subjected to CCK8 assay to detect the absorbance at 450 nm.

Transfected/transduced U251 or U87 cells (2×10³ cells/well) were seeded in 96-well plates and incubated for 24, 48, 72 and 96 h. Subsequently, 20 µl Cell Counting Kit-8 (CCK8) solution (Dojindo Molecular Technologies, Inc.) was added into each well and incubated at 37°C for 2 h, according to the manufacturer’s protocols. The absorbance was measured at 450 nm using a Microplate Autoreader (Thermo Fisher Scientific, Inc.).