Mia paca 2 cells

Human cervical cancer-derived HeLa cells, human pancreas carcinoma-derived MIA PaCa-2 cells, and human pancreas adenocarcinoma-derived BxPC-3 cells were purchased from the Riken BRC Cell Bank (Ibaraki, Japan). The human epidermoid carcinoma-derived A431 cells were purchased from the American Type Culture Collection (Manassas, VA, USA). The cells were cultured in minimum essential medium α (α-MEM; Gibco, Life Technologies Corporation, Grand Island, NY, USA) containing 10% heat-inactivated foetal bovine serum (FBS; Gibco, Life Technologies Corporation) (HeLa cells), minimum essential medium (MEM) (Gibco, Life Technologies Corporation) containing 10% heat-inactivated FBS (Gibco, Life Technologies Corporation) (A431 cells), Eagle’s minimum essential medium (EMEM) (Wako, Osaka, Japan) containing 10% FBS (HyClone Laboratories, South Logan, UT, USA), 0.1 mM MEM non-essential amino acids (Gibco), penicillin-streptomycin (50 units/ml and 50 μg/ml) (Sigma-Aldrich, St. Louis, MO, USA) (MIA PaCa-2 cells), and RPMI1640 (Gibco, Life Technologies Corporation) containing 10% FBS (HyClone), and penicillin-streptomycin (50 units/ml and 50 μg/ml) (Sigma-Aldrich) (BxPC-3 cells). Cells were grown on 100-mm dishes and incubated at 37 °C under 5% CO₂.

MiaPaCa-2 cells, obtained from the Bioresource Collection and Research Center, Hsinchu, Taiwan, and the Detroit 551 cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM). All cells were grown in a humidified CO₂ incubator at 37°C. Cell viability was determined by the MTS assay. In brief, cells were cultured (2,500–3,000 cells/well) in flat bottomed 96-well plates for 24 h. Serial-diluted conjugates were added to the cell media and incubated for 72 h. Culture medium was removed after 72 h followed by the addition of 100 μl of MTS and PMS mixture solution. In a humidified incubator at 37°C with 5% CO₂, cells were then incubated for another 1.5 h for viable cells to convert the tetrazolium salt into formazan. The conversion to formazan was monitored at the 490-nm absorbance by a BioTek PowerWave-X Absorbance Microplate Reader. In this MTS assay, the recorded data were normalized to DMSO-treated and background controls. Using GraphPad Prism version 4 software, the IC₅₀ value was the amount of each conjugate that caused a 50% reduction in cell viability in comparison to DMSO-treated controls.