168 1000 model 680

The in vitro cell proliferation assay was performed as described previously.^{31 (link)} A total of 10³ MXD1 (OriGene Technologies, Beijing, China) or si-MXD1 (Invitrogen) cells or cells transiently transfected with the control vector were used for the assays in 200 μl of complete medium. For proliferation of pre-miR-19a/b and miR-19a/b-inh, SGC7901 cells were used for transfection using pre-miRNAs, miRNA inhibitors and their negative controls, respectively. Twenty four hour later, cells were resuspended and 10³ cells were used for the assays in 200 μl of complete medium. The cultures were assayed each day and read at a 490 nm absorbance (A490) on a micro-plate reader (168–1000 Model 680, Bio-Rad Laboratories, Inc.). Each experiment was performed in triplicate and repeated three times.

Cell Counting Kit‐8 (CCK‐8; Biotool, Houston, TX, USA) was used to measure cell proliferation. Cells (2 × 10³/well) were seeded into 96‐well plates containing complete DMEM (100 μL) in triplicate for each condition and were maintained in an incubator at 37°C with 5% CO₂. Then CCK‐8 solution (10 μL) was added to each well and incubated for 4 h. Optical density values were measured with a water‐soluble tetrazolium salt assay using microplate computer software (Bio‐Rad Laboratories, Hercules, CA, USA) according to the protocol of the CCK‐8 assay kit (Biotool). The absorbance at 450 nM (A450) was read on a microplate reader (168–1000 Model 680; Bio‐Rad Laboratories), and proliferation curves were plotted.