The acute monocytic leukemia cell line THP1 (ATCC® TIB–202™), promyelocytic leukemia cell line HL60 (ATCC® CCL–240™), and the multiple myeloma cell line RPMI8226 (ATCC® CCL–155™) were obtained from the American Type Culture Collection (ATCC, Rockville, MD, USA). All the investigated cells were cultured in RPMI 1640 medium supplemented with 10% fetal bovine serum, penicillin (10 U/mL), and streptomycin (50 μg/mL) in standard conditions: 37 °C, 100% humidity, and an atmosphere of 5% CO2 and 95% air. Cell viability was systematically controlled using 0.4% trypan blue. In all experiments, cells in a logarithmic growth phase were used when their viability was above 95%.
One concentration of the drugs for each cell line was chosen for the study: THP1: 0.3 µM; HL60: 0.7 µM; RPMI8226: 3 µM. These were the same as in previous studies evaluating the biological properties of melphalan analogs [14 (link),15 (link)]. In all experiments, cells were treated for 4 h, 24 h, and 48 h. Simultaneously, control cultures were incubated similarly without further treatment. For the immunostaining assay, a positive control from irradiated cells was prepared. The cells were irradiated with 1 Gy of ionizing radiation by an ISOVOLT Titan X-ray generator (GE, Ahrensburg, Germany) [51 (link),52 (link)].
One concentration of the drugs for each cell line was chosen for the study: THP1: 0.3 µM; HL60: 0.7 µM; RPMI8226: 3 µM. These were the same as in previous studies evaluating the biological properties of melphalan analogs [14 (link),15 (link)]. In all experiments, cells were treated for 4 h, 24 h, and 48 h. Simultaneously, control cultures were incubated similarly without further treatment. For the immunostaining assay, a positive control from irradiated cells was prepared. The cells were irradiated with 1 Gy of ionizing radiation by an ISOVOLT Titan X-ray generator (GE, Ahrensburg, Germany) [51 (link),52 (link)].
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