Mcdb 105

Human ovarian cancer cell line OVCAR5 (female) was obtained from American Type Culture Collection. A2780 (female) was acquired from European Collection of Cell Cultures. OVCA433 (female) ovarian serous adenocarcinoma cell line was a generous gift from Dr. Marcin Iwanicki (Stevens Institute of Technology, Department of Chemistry and Chemical Biology, NJ USA). OVCAR5 and A2780 were grown in RPMI-1640 medium (Corning, catalogue No. 10-040-CM). OVCA433 (Iwanicki et al., 2016)^{39 (link)} were cultured in 1:1 mixture of MCDB 105 (Cell Applications, catalogue No. 117–500) medium and Medium 199 (Gibco, catalogue No. 11150059). All media were supplemented with fetal bovine serum (FBS; Gemini Bio-products, catalogue No. 900–208) and penicillin-streptomycin solution (Corning, catalogue No. 30-002-Cl) at the final concentrations of 10% and 1%, respectively, unless stated otherwise. Cells were kept in a humidified incubator with 5% CO₂ at 37 °C. All cells were tested for mycoplasma infection (MycoAlert Mycoplasma Detection Kit, Lonza, catalogue No. LT07-218) and found to be negative. Cells were counted by means of Bright-Line^™ Hemacytometer (Sigma, catalogue No. Z359629).

Ovarian cancer cell lines OVCAR-3 (HTB-161, ATCC, Manassas, VA, USA), SKOV-3 (HTB-77, ATCC) and OV90 (CRL-11732, ATCC), astrocytoma cell line E98 [67 (link)] and primary skin fibroblasts C5120 [46 (link)] were cultured in a humidified incubator at 37 °C and 5% CO₂. SKOV-3 and E98 cells were cultured in DMEM (Gibco) with 10% fetal calf serum (FCS). OVCAR-3 cells were cultured in RPMI (Gibco) with 20% FCS. Following recommendation by ATCC, OV90 cells were cultured in MCDB 105 (Cell Applications) and Medium 199 (Sigma-Aldrich) (1:1, v:v) with 15% FCS. C5120 cells were cultured in Medium 199 with 10% FCS.
Tumor cell spheroids of OV90 and SKOV-3 cells were formed by the hanging drop method as described earlier [68 (link)]. Cells were detached with trypsin/EDTA and resuspended in complete culture medium containing 1.2 mg/mL methylcellulose (Sigma-Aldrich) with 200 U/mL penicillin/streptomycin (Sigma-Aldrich). Then, 15,000 cells were seeded in 30 µL drops hanging from the inverted lid of a 140 × 20.6 mm petri dish (VWR international). Spheroids were used for experiments after 48 or 72 h.

McCoy’s 5A (#16600-082), Medium 199 (#11150-059), RPMI1640 (#11875-085), DMEM/F12 (#11320-033), 1% Penn Strep (Penicillin/Streptomycin; P/S, #15140122), MEM Non-Essential Amino Acids (NEAA) Solution (100X, #11140-050), DPBS (#14040-117), and Trypsin-EDTA (0.25%, #25200-056) were purchased from Gibco (Life Technologies, Grand Island, NY). MCDB105 (#117-500) was purchased from Cell Applications Inc. (San Diego, CA). Alpha-MEM was obtained from Irvine Scientific (Irvine, CA) OCMI-E was purchased from University of Miami (Live Tumor Culture Core at Sylvester Comprehensive Cancer Center, University of Miami Miller School of Medicine, Miami, FL). Fetal bovine serum (FBS, #F2442) and Cholera Toxin from Vibrio cholerae (#C8052) were purchased from Sigma-Aldrich (St. Louis, MO). Dimethyl Sulfoxide (DMSO, #CAS 67-68-5) was purchased from Thermo Fisher Scientific (Waltham, MA).