Annexin 5 propidium iodide apoptosis staining kit

WM cell lines and their corresponding ibrutinib-resistant clones, developed in our laboratory, were used in experiments. All cell lines were cultured in RPMI-1640 containing 10% fetal bovine serum, penicillin (100 U/ml) and streptomycin (100 μg/ml). Cell viability was always maintained at >90% and was measured by trypan blue exclusion assay using ViCell-XR viability counter (Beckman-Coulter, Indianapolis, IN, USA). RPMI, penicillin, streptomycin, tetramethylrhodamine, methyl ester (TMRM) and fetal bovine serum were purchased from Life technologies (Carlsbad, CA, USA). Ibrutinib, MK2206 and ABT-199 (venetoclax) were purchased from Sellekhem (Houston, TX, USA). Annexin-V/Propidium Iodide Apoptosis Staining Kit was purchased from BD Biosciences (San Jose, CA, USA).

WM cell lines and derived bortezomib-resistant (BR) suclones,^{24 (link), 25 (link)} as well as ibrutinib-resistant (IR) subclones,^{26 , 27 (link)} were used in experiments, as reported previously.^{23 (link), 25 (link)} Primary WM patient cells (CD19+/CD138+ sorted) were obtained from the Predolin Biobank (Mayo Clinic, Rochester MN, USA) after approval by the Mayo Clinic Institutional Review Board. Heparinized peripheral blood from healthy human donors was obtained and peripheral blood mononuclear cells (PBMCs) were extracted, as described previously.^{28 (link)} All cells were cultured according to conditions previously described by us.^{23 (link)} VLX1570 and b-AP15 were provided as gifts from Vivolux AB (Uppsala, Sweden). RPMI, penicillin, streptomycin, tetramethylrhodamine, methyl ester (TMRM) and fetal bovine serum were purchased from Life Technologies (Carlsbad, CA, USA). Ibrutinib and bortezomib were purchased from Sellekhem (Houston, TX, USA). Annexin-V/Propidium Iodide Apoptosis Staining Kit was purchased from BD Biosciences (San Jose, CA, USA).