Celltiter glo ctg luminescent cell viability assay

The primary screen was performed in 384-well format using the CellTiter Glo (CTG) luminescent cell viability assay (Promega) as previously described²⁷ . 3630 compounds from the St. Jude proprietary molecular glue library were screened against 9 human cancer cell lines (HD-MB03, MB004, MB002, MHH-CALL4, MOLM-13, TF-1, HEL, OCI-AML3, AML193). Each cell line was cultured in the complete medium recommended by the vendor and seeded in Corning 8804 BC white 384-well assay plates at densities of 1000, 1000, 1500, 7500, 1250, 156, 625, 1250, 1250 cells per well for HD-MB03, MB004, MB002, MHH-CALL4, MOLM-13, TF-1, HEL, OCI-AML, AML193, respectively. After overnight incubation at 37 °C in a humidified 5% CO₂ incubator, cells were treated with compounds in dose–response format using a Pintool on a Biomek FX^P Laboratory Automation Workstation (Beckman Coulter). After 72 h of incubation, cell proliferation was assessed using a CellTiter-Glo (CTG) luminescent cell viability assay (Promega) according to the manufacturer’s instructions. Luminescence signal was measured using an EnVision plate reader (PerkinElmer).

Growth-inhibitory effects of varying concentrations of 6-thio-dG in combination with etoposide (VP-16, Selleckchem, USA), doxorubicin (Merck, Germany) or ceritinib (LDK378, Selleckchem, USA) on neuroblastoma cell lines with TERT rearrangements (CLB-GA, GI-ME-N), high TERT expression (SH-SY5Y) or MYCN amplification (BE(2)-C, IMR-32, Kelly, LS, TR-14) were determined after 96 h of substance incubation using CellTiter-Glo®. A semi-automated substance screening protocol was established on a Biomek 4000 Automated Workstation (Beckman Coulter, Brea, CA, USA). Single or combined compounds in nine different concentrations and DMSO as control were added in triplicates 24 h after the cells were seeded in a 96-well plate at a density of 3000 cells/well. Cell viability was measured after 96 h of substance incubation using a CellTiter-Glo® (CTG) Luminescent Cell Viability Assay (Promega, Madison, WI, USA). In brief, 100 µl CTG-reagent was added to the wells and incubated for 30 min at room temperature. Signals were measured using a SpectraMax® i3x Multi-Mode microplate reader (Molecular Devices, San Jose, CA, USA). Drug synergy was calculated according to the Chou-Talalay combination index method [17 (link), 18 (link)].

Cell proliferation assay was performed using methods reported in the previous article. Cell proliferation was measured using the CellTiter-Glo^® (CTG) Luminescent Cell Viability Assay (Promega, WI, USA). Cells were seeded in 96-well plates at 15,000 cells per well in detection medium containing 0.5% fetal bovine serum and placed in an incubator at 37℃ and 5% CO₂ overnight. On the following day, iruplinalkib was serially diluted with DMSO, further diluted in assay medium, and added to the designated wells for serial dilution of iruplinalkib to the target gradient concentration. After the cells were incubated for 24 h, the number of viable cells at each concentration was determined by CTG assay, and the half-maximal inhibition concentration (IC₅₀) values were calculated by concentration-response curve fitting utilizing a four-parameter logistic regression model.