Evaluating Colon Cancer Cell Viability

Colonic tumor tissues were received from consented patients following Institutional Review Board (IRB) approval at the Norris Comprehensive Cancer Center of the University of Southern California, Los Angeles, CA, USA. Colonic tumor specimens were isolated from colon cancer patients and processed mechanically and enzymatically to obtain colon cancer cells without contaminating non-cancer cells as previously described^{37 (link)}. Patient-derived organoids (PDOs) were generated from these colon cancer cells as described in our recent publication^{6 (link)} and treated with B32B3 and Taz individually or in combination for 5 days. At the end of treatment, 100 μl of CellTiter-Glo 3D solution (Promega, WI, USA) was added to each well, and cell viability was analyzed by measuring luminescence with BioTek Synergy Neo2 Multi-Mode Reader (Agilent Technologies, CA, USA). Images of PDOs were also acquired 0 and 5 days after inhibitor treatments using the Operetta CLS high-content analysis system (PerkinElmer, MA, USA) and displayed with maximum intensity projections of 24 z-stacks ranging from 10–470 µm in increments of 20 µm.