HGSOC cell lines were cultured in 100 mm petri dishes (Sarstedt Inc., Nümbrecht, Germany) in OSE medium (WISENT Inc., St-Bruno, QC, Canada) supplemented with 10% fetal bovine serum (WISENT Inc.), 0.5 µg/mL amphotericin B (WISENT Inc.) and 50 µg/mL gentamycin sulfate (WISENT Inc.) (complete OSE medium). Plates were maintained at 37 °C in low oxygen conditions (7% O2 and 5% CO2). Cells were passaged at near confluence by trypsin 0.05% (WISENT Inc.) digestion. Cultures were discarded before the 20th passage, after which a fresh batch of cells was thawed for further experiments. For resistant cell lines, passages were counted from when stable resistance was confirmed.
Ose medium
Manufactured by Wisent
Sourced in Canada
OSE medium is a cell culture medium formulated for the growth and maintenance of embryonic stem cells. It provides the necessary nutrients and growth factors to support the cultivation of undifferentiated stem cells.
Automatically generated - may contain errors
Lab products found in correlation
Amphotericin b, by Wisent (7 mentions)
Gentamicin, by Thermo Fisher Scientific (5 mentions)
Fetal bovine serum (fbs), by Wisent (4 mentions)
Gentamycin sulfate, by Wisent (2 mentions)
100 mm petri dishes, by Sarstedt (1 mentions)
Foetal bovine serum (fbs), by Wisent (1 mentions)
Trizol, by Thermo Fisher Scientific (1 mentions)
Dmem f12 50 50 medium, by Wisent (1 mentions)
Dmem f12, by Wisent (1 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (1 mentions)
L glutamine, by Thermo Fisher Scientific (1 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (1 mentions)
Amphotericin b, by Thermo Fisher Scientific (1 mentions)
Gentamicin, by Merck Group (1 mentions)
Du145, by American Type Culture Collection (1 mentions)
Skov3, by American Type Culture Collection (1 mentions)
Rpmi 1640 medium, by Wisent (1 mentions)
Lncap, by American Type Culture Collection (1 mentions)
Mcf 7, by American Type Culture Collection (1 mentions)
Skov3, by Wisent (1 mentions)
10 protocols using ose medium
1
Culturing HGSOC Cell Lines
2
Establishment of Ovarian Cancer Cell Lines
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In total, six cell lines were derived from samples from six patients, 866, 2978, 3041, 3291, 4453 and 4485. All cell lines were maintained in a low oxygen condition of 7% O2, and 5% CO2 and grown in complete OSE medium, which includes OSE medium (Wisent, St-Bruno, QC), 10% FBS, 0.5 μg/mL amphotericin B (Wisent) and 50 μg/mL gentamicin (Gibco®, Life Technologies Inc., Burlington, ON). The solid ovarian tumor (TOV)-derived cell lines (TOV2978G, TOV3041G, TOV3291G) were established using the scrape method as previously described [11 (link), 76 (link)]. Briefly, tumor tissue was scraped into a 100 mm plate with complete OSE medium and maintained for 40 days with the medium replaced weekly. Cells were passaged at near confluence, and were considered immortal when passaged over 50 times. The OV cell lines (OV866(2), OV4453, OV4485) were established from the cellular fraction of ascites collected by centrifugation [11 (link), 76 (link)]. The cell lines derived from ascites cells were maintained as above for the TOV derived cell lines. Although to date each cell line has reached at least 90-100 passages, most assays were conducted on cell lines between passage 60 and 80.
3
Characterization of HGSC Cell Lines
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All 18 cell lines used in our study were derived from HGSC solid tumors or peritoneal ascites and were described in publications from our group. Their characteristics are summarized in Table S1 [58 (link),59 (link),60 (link)]. Cell lines were cultivated at 37 °C in hypoxic condition of 7% O2, and 5% CO2 and grown in OSE medium (Wisent, St.-Bruno, QC, Canada) supplemented with 10% Foetal Bovine Serum, 0.5 μg/mL amphotericin B (Wisent), and 50 μg/mL gentamicin (Gibco®, Life Technologies Inc., Waltham, MA, USA).
4
Ovarian Cell Line Culture Protocol
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The ovarian adenocarcinoma SKOV3ip1 and the ovarian immortalized INOF cell line were grown in DMEM/F12 (50/50) medium and OSE medium (Wisent), respectively. The medium in both cases was supplemented with 10% fetal bovine serum and 2 mM L-glutamine. Cell propagation and passaging were as recommended by ATCC (American Type Culture Collection). Cells were trypsinized and collected in 5 × 106 pellets, resuspended in 700 µL TRIzol (Ambion) and kept at −80°C until RNA extraction.
5
Detailed Characterization of Human EOC Cell Lines
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The human EOC cell lines [TOV81D, TOV112D, TOV1946, OV866(2), OV1946, TOV2295(R), OV4485] were derived in our laboratory from patients’ tumors (TOV) or ascites (OV) and characterized in detail [34 (link)–38 (link)]. All EOC cell lines were maintained in a low oxygen condition of 7% O2 and 5% CO2 and grown in OSE medium (Wisent, Montreal, QC). The human retinal epithelial cell line ARPE was purchased from American Type Culture Collection (ARPE-19, #CRL2302) and maintained in DMEM-F12 (Wisent). All media were supplemented with 10% FBS (Wisent), 0.5 μg/mL amphotericin B (Wisent), and 50 μg/mL gentamicin (Life Technologies Inc). Antibodies, chemicals, oligonucleotides, plasmids, and commercial assays used in this study are indicated in Supplementary Table S1 .
6
Establishment of Ovarian Cell Lines
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The HGSC cell line OV-4453 was established from the ascites of a patient whereas TOV-1369TR and TOV-1369 (2) were established from a HGS ovarian tumor of a patient before (TOV-1369TR) and after (TOV-1369 (2)) chemotherapy treatment63 (link),64 (link). All cell lines were maintained in ovarian surface epithelium (OSE) medium (Wisent, QC, Canada) supplemented with 10% [v/v] fetal bovine serum.
7
Human Ovarian Cancer Cell Lines
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The 18 human HGS EOC cell lines used (OV90, OV866(2), TOV1369, OV1369(R2), TOV1946, OV1946, TOV2223G, OV2295, OV2295(R2), TOV2295(R), TOV2978G, TOV3041G, TOV3291G, TOV3133G, TOV3133D, OV3133(R), OV4453, OV4485) were derived in our laboratory from patients’ tumors (TOV) or ascites (OV) [22 (link)–25 (link)]. All cell lines were maintained in a low oxygen condition of 7% O2 and 5% CO2 and grown in complete OSE medium, which contains OSE medium (Wisent, Montreal, QC) with 10% FBS (Wisent), 0.5 μg/mL amphotericin B (Wisent) and 50 μg/mL gentamicin (Life Technologies Inc., Burlington, ON).
8
Establishment and Characterization of Ovarian Cancer Cell Lines
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VOA-3723, VOA-4627, and VOA-6406 (RRID:CVCL_VQ52) cells were established by the Mark Carey lab (8, 9 ). The iOvCa241-PAR and iOvCa241-RES (selected under 20 nmol/L trametinib) cells were provided by the Gabriel DiMattia lab. All cells were maintained in DMEM (Gibco) supplemented with 10% FBS and 100 μg/mL penicillin/streptomycin and 2 mmol/L L-glutamine (Gibco). For primary ascites cultures, 25-mL fresh ascites was combined with 25-mL OSE medium in a T175 flask until confluency. The OSE medium (Wisent Bio Products) was supplemented with 10% FBS, 2.5 μg/mL amphotericin B (Thermo Fisher), and 50 μg/mL gentamicin (Sigma; ref. 10 (link)). For the ascites cultures, written informed consent was obtained in accordance with the declaration of Helsinki, and the use was approved by our institutional review board. The ascites cultures proliferated only a few passages. Further authentication of the obtained cell lines was not performed. RAS mutation status of the VOA-6406 and IOvCa241 cell lines was confirmed by Sanger sequencing. All cultures were frequently checked for Mycoplasma using a qPCR-based method.
9
Cell Culture Protocols for Cancer Research
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Jurkat T lymphoma cells were kindly provided by Dr. Lapointe Réjean (CRCHUM), while MCF-7, SKOV3, and all PC cell lines (22Rv1, LNCaP, DU145, and PC3) were obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA). Jurkat T lymphoma cell and PC cells were maintained in RPMI 1640 medium (Wisent Inc., St-Bruno, QC, Canada), MCF-7 was grown in DMEM medium (Wisent Inc.), and SKOV3 in OSE medium (Wisent Inc.). All culture media were supplemented with 10% fetal bovine serum (FBS) (Gibco®, Thermo Fisher Scientific, Waltham, MA, USA), 0.454 μg/mL of amphotericin B (Wisent Inc.), and 90 μg/mL gentamycin sulfate (Wisent Inc.).
10
Characterization of HGSOC Cell Lines
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The four human HGSOC cell lines used, OV1369(R2), OV90, OV4453, and OV1946, were derived in our laboratory from the ascites of patients diagnosed with HGSOC and have been extensively characterized23 (link)–26 (link). All cell lines were maintained in a low oxygen condition (7% O2 and 5% CO2) and grown in OSE medium (Wisent, Montreal, QC) with 10% fetal bovine serum (FBS) (Wisent), 0.5 μg ml−1 amphotericin B (Wisent) and 50 μg ml−1 gentamicin (Life Technologies Inc., Burlington, ON). The MDA-MB-231 breast cancer cell line was a gift from the laboratory of Dr. John Stagg (CRCHUM, Canada) which was purchased from ATCC. It was maintained in DMEM (Wisent) with 10% FBS (Wisent), 0.5 μg ml−1 amphotericin B (Wisent) and 50 μg ml−1 gentamicin (Life Technologies Inc.). All HGSOC cell lines were authenticated in 2017 using short tandem repeat (STR) profiling by the McGill University Genome Center (Montreal, Canada). All cell lines were tested negative for mycoplasma with IDEXX BioAnalytics (Columbia, MO65201).
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