Sulforhodamine b srb reagent

PD173074 (Sigma-P2499, HPLC ≥ 96%) and olaparib (AZD2281/KU0059436, #S1060, HPLC ≥ 99.7%) were purchased from Sigma-Aldrich Co. (St. Louis, MO, USA) and Selleck Chemicals (Antibody International Inc. Jhubei City, Hsinchu County, Taiwan), respectively. Stock solutions (1 mM) of each drug were prepared by dissolution in phosphate-buffered saline (PBS) and stored in a dark room at −20 °C. PBS, dimethyl sulfoxide (DMSO), sulforhodamine B (SRB) reagent, trypsin/ethylenediaminetetraacetic acid, Tris aminomethane (Tris) base, and acetic acid were purchased from Sigma-Aldrich Co. (St. Louis, MO, USA). Dulbecco’s modified Eagle’s medium (DMEM) was purchased from Invitrogen (Invitrogen Life Technologies, Carlsbad, CA, USA).

BX-795 hydroxide (#SML0694, HPLC ≥ 98%) was purchased from Sigma Aldrich Co. (St. Louis, MO, USA). Stock solutions of 1 mM were dissolved in dimethyl sulfoxide (DMSO) at 15 mg/mL, and stored in dark room at −20 °C. Phosphate buffered saline (PBS, #P7059), dimethyl sulfoxide (DMSO, #D2650), sulforhodamine B (SRB) reagent (#230162), trypsin/ethylenediaminetetraacetic acid (Trypsin-EDTA, #T4049) solution, trisaminomethane (Tris) base (#93352) and acetic acid (#695092) were purchased from Sigma Aldrich Co. (St. Louis, MO, USA), while Gibco^TM Dulbecco’s modified Eagle’s medium (DMEM) was purchased from Invitrogen (#11966025, Invitrogen Life Technologies, Carlsbad, CA, USA).

The cell viability assay was performed using sulforhodamine B (SRB) reagent (Sigma-Aldrich, Taipei, Taiwan), as described previously [88 (link)]. Briefly, DLD-1 and HCT116 cells were harvested at 90–95% confluency, and 8000 cells/well were seeded in 96-well plates for a period of 24 h, followed by dose-dependent concentration treatment of LCC-21. After 48 h of treatment, 10% trichloroacetic acid (TCA) was added to cells and they were stored at 4 °C for one hour. Cells were further washed twice with distilled water and stained with 0.4% SRB, then stored for 30 min at room temperature. Excess stain was removed from the plates by washing with 1% acetic acid twice. The plates were air-dried overnight. The protein-bound stain was solubilized with a 20 mM Tris-buffer solution for 15 min on an orbital shaker. The absorbance was measured with a microplate reader at a wavelength of 560 nm (Molecular Devices, Sunnyvale, CA, USA).