All tissue samples were obtained from patients treated at The First Affiliated Hospital of Zhengzhou University. Each patient signed an informed consent form, and this study was approved by the First Affiliated Hospital of Zhengzhou University Institutional Review Board. Tissues samples were collected from OC patients with unilateral ovarian invasion and normal contralateral ovary, including 17 cases of high-grade serous carcinoma, 2 cases of mucinous cystadenoma and 1 case of mucinous carcinoma. Human OC cell lines (A2780, SKOV3, OVCAR3) and a normal ovary cell line (IOSE80) were purchased from the Cell Bank of Chinese Academy of Sciences (Shanghai, China). All cell lines were authenticated using the short tandem repeat (STR) profiling test. Then, IOSE80, A2780, OVCAR3 cells were cultured in RPMI-1640 Medium (Biological Industries, USA) supplemented with 10% fetal bovine serum (FBS, Biological Industries, USA), 100 units/ml penicillin, and 100 μg/ml streptomycin (Biological Industries, USA). SKOV3 cell was maintained in McCoy’s 5A Medium (Biological Industries, USA) supplemented with 10% FBS, 100 U/ml penicillin, and 100 μg/ml streptomycin.
Mccoy s 5a medium
Manufactured by Sartorius
Sourced in Israel, United States
McCoy's 5A medium is a culture medium formulated for the growth of a variety of cell types. It provides the necessary nutrients and growth factors to support the proliferation of cells in vitro.
Automatically generated - may contain errors
Lab products found in correlation
Fetal bovine serum (fbs), by Sartorius (10 mentions)
Rpmi 1640 medium, by Sartorius (6 mentions)
Penicillin streptomycin, by Sartorius (5 mentions)
Lipofectamine 2000, by Thermo Fisher Scientific (4 mentions)
Hct116, by Sartorius (4 mentions)
Dulbecco modified eagle medium (dmem), by Sartorius (4 mentions)
Penicillin, by Sartorius (3 mentions)
Streptomycin, by Sartorius (3 mentions)
L glutamine, by Sartorius (3 mentions)
Dulbecco s modified eagle medium, by Sartorius (3 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (3 mentions)
A2780, by Sartorius (2 mentions)
Ovcar3, by Sartorius (2 mentions)
Rcc jf, by Corning (2 mentions)
Rcc 23, by Thermo Fisher Scientific (2 mentions)
Caki 1, by Sartorius (2 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (2 mentions)
Caki 1, by American Type Culture Collection (2 mentions)
Rpmi 1640 medium, by Corning (2 mentions)
Skov3, by Sartorius (2 mentions)
31 protocols using mccoy s 5a medium
1
Ovarian Cancer Tissue Samples and Cell Lines
2
Characterization of HCT116 Cell Lines
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HCT116WT and HCT116p53null cell lines were provided by Dr Bert Vogelstein at The John Hopkins University Medical School28 and grown in McCoy’s 5A medium (Biological Industries) with 10% FBS (Biological Industries) and 1% penicillin‐streptomycin. Mycoplasma contamination was routinely tested using the Mycoplasma Detection Kit‐QuickTest (Biotool). All cells were cultured in a humidified atmosphere of 5% CO2 at 37°C. Transfection was performed using Lipofectamine 2000 (Invitrogen Life Technologies) according to the manufacturer’s protocol. For gene‐silencing experiments, to eliminate untransfected cells, 24 hours after transfection, transfected cells were selected by using puromycin (final concentration: 1.2 µg/mL) for 36 hours.
For overexpression experiments, cells were transfected with 2 µg of indicated overexpression vector. Twenty‐four hours later, mRNA and protein samples were collected and subjected for further analysis.
For double‐silencing experiments, cells were transfected with 1 µg of indicated shRNA expression vector. Cells were subjected to puromycin selection to eliminate untransfected cells. mRNA and protein were collected 36 hours after puromycin selection. For NeuroD1‐silenced HCT116 cells, NeuroD1, p53‐double silenced HCT116 cells or control stable cell lines, cells were transfected with the indicated vectors (total 2 µg) and selected with puromycin.
For overexpression experiments, cells were transfected with 2 µg of indicated overexpression vector. Twenty‐four hours later, mRNA and protein samples were collected and subjected for further analysis.
For double‐silencing experiments, cells were transfected with 1 µg of indicated shRNA expression vector. Cells were subjected to puromycin selection to eliminate untransfected cells. mRNA and protein were collected 36 hours after puromycin selection. For NeuroD1‐silenced HCT116 cells, NeuroD1, p53‐double silenced HCT116 cells or control stable cell lines, cells were transfected with the indicated vectors (total 2 µg) and selected with puromycin.
3
Gadolinium Nanoparticles Induce HT29 Cell Apoptosis
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All of the chemicals were analytical grade reagents and without further purification. GdNO3·6H2O and catalase (CAT) from bovine live (≥40,000 units/mg protein) were purchased from J&K Scientific Ltd. (Beijing, China). Dopamine hydrochloride (DA), DCFH-DA, DAPI, Hoechst33342, and PI were supplied by Shanghai Aladdin Biochemical Technology Co., Ltd. (Shanghai, China). Na2WO4·2H2O was obtained from the Guoyao Chemical Research Institute (Shenyang, China). The HT29 cell line was obtained from the Cell Culture Center of the Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences. 3-[4,5-di-methylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) was purchased from Sigma-Aldrich (Shanghai, China). Annexin V-FITC apoptosis detection kit and Goat anti rabbit IgG-FITC were supplied by Absin Biotechnology Co., Ltd. (Shanghai, China). HIF-1α antibody and γ-H2AX antibody were purchased from Cell Signaling Technology, Inc. (Danfoss, CA, USA). McCoy’s 5A medium was obtained from Biological Industries (Shanghai, China). Fetal bovine serum was purchased from Sigma-Aldrich (Shanghai, China).
4
Culturing and Transfecting Renal Cancer Cell Lines
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Four strains of ccRCC cell lines (including 786-O, RCC-JF, RCC-23, and Caki-1) and human renal tubular epithelial cell HK-2 were obtained from American Type Culture Collection (Manassas, VA, USA). The 786-O, RCC-JF, and HK-2 were incubated in RPMI 1640 medium (Corning Inc., Corning, NW, USA), RCC-23 was incubated in Dulbecco's modified Eagle's medium (DMEM) (Gibco, Amarillo, TX, USA), and Caki-1 was cultured with McCoy's 5A medium (Biological Industries, Israel). All media were supplemented with 10% fetal bovine serum (Gibco, Thermo Fisher Scientific, MA, USA), and cells were incubated at 37°C in 5% CO2. All small interfering RNAs (siRNAs) targeting human lncRNA LINC00342 (si-LINC00342-1, si-LINC00342-2) or scrambled negative control (si-NC) were designed and synthesized by GenePharma (Shanghai, China; Table 2 ), transfected into the cell lines with Lipofectamine 2000 (Invitrogen, Carlsbad, CA, USA), according to the instructions of the manufacturer.
5
Culturing Human Ovarian Cancer and Monocyte Cells
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Two human ovarian cancer cell lines (SKOV3 and A2780) and a human monocyte line THP-1 were purchased from Procell Life Science & Technology. SKOV3 cells were cultured in McCoy’s 5 A medium (Biological Industries) containing with 10% fetal bovine serum (FBS, Biological Industries) and 1% penicillin/streptomycin (Beyotime). A2780 cells were cultured in RPMI 1640 (Biologial Industries), containing with 10% FBS and 1% penicillin/streptomycin. THP-1 cells were maintained in RPMI 1640 medium containing with 10% FBS. All cells were cultured in an incubator with 5% CO2 at 37 °C.
6
Cell Line Culturing for Epigenetic Studies
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HCT116 parental colorectal cell line (HD PAR-033), as well as KO (DNMT3B-/-, HD R02-023) and DKO (DNMT1exons3-5/exons3-5; DNMT3B-/-, HD R02-022) were obtained from Horizon Discovery Ltd., Cambridge, UK. A549 cells were obtained from the NCI-60 cell panel, NCI-H1299 cells were purchased from ATCC (ATCC® CRL-5803™) and WI38-hTERT embryonic lung fibroblasts (WI38) were generated as described in Milyavsky et al.21 (link). All Cells were cultured on 100 × 20 mm culture dishes (Corning, 353063) in heat-inactivated medium and split at a ratio of 1:10 every 2–3 days using 0.05% trypsin-EDTA solution C (Biological Industries, Israel; 03-053-1B). McCoy’s 5A medium (Biological Industries, Israel; 01-075-1A) was used for HCT116 WT, KO and DKO, DMEM medium (Dulbecco’s Modified Eagle Medium, Biological Industries, Israel; 01-050-1A) was used for WI38, DMEM/F-12 (HAM) medium (Biological Industries, Israel; 01-170-1A)) was used for A549 cells and RPMI medium (Biological Industries, Israel; 01-100-1A) was used for NCI-H1299 cells. All media were supplemented with 10% Fetal Bovine Serum (GibcoTm FBS, 10270-106), 0.4% Penicillin-Streptomycin (Biological Industries, Israel; 01-031-1C) and 1% L-glutamine (Biological Industries, Israel; 01-020-1A). Modified medium was filtered through a 0.22-μm filter (Corning, 430769) prior to culture.
7
Cell Culture of Human Cancer Cells
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The human CRC cell lines HCT116, HT29 and human embryonic kidney 293T (293T) cells were purchased from Shanghai Zhong Qiao Xin Zhou Biotechnology Co., Ltd. The HCT116 and HT29 cell lines were cultured in McCoy's 5A Medium (Biological Industries), and the 293T in EMDM Medium supplemented with 10% fetal bovine serum (Biological Industries) at 37°C in 5% CO2.
8
Cell Culture Protocol for HEK293T, COS7, and HCT116
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HEK293T (internal stock) and COS7 (internal stock) cells were cultured in complete DMEM (Biological Industries, Israel), and HCT116 (kind gift from Prof. Moshe Oren, Weizmann Institute, Israel) cells were cultured in McCoy’s 5A medium (Biological Industries). Culture media were supplemented with 10% heat-inactivated fetal bovine serum (FBS), 2 mM L-glutamine, 1 mM sodium pyruvate, 100 U/mL penicillin G sodium, and 0.1 mg/mL streptomycin sulfate. Cultures were maintained at 37 °C in a humidified atmosphere containing 5% CO2. All cell lines were confirmed to be mycoplasma-free using the EZ-PCR Mycoplasma Detection Kit (Biological Industries).
9
Culturing Ovarian Cancer Cell Lines
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The SKOV3, A2780 and OVCAR-3 human epithelial ovarian cancer cell lines were obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA). Their TP53 status were null (SKOV3), wild-type (A2780) and mutant (OVCAR-3). The SKOV3 cells were maintained in McCoy's 5A medium (Biological Industries, Beit Haemek, Israel), A2780 cells and OVCAR-3 cells were cultured in RPMI 1640 medium (Biological Industries), both all supplemented with 10% fetal bovine serum and 1% penicillin/streptomycin. Cells were maintained at 37°C with an atmosphere of 5% CO2, following standard protocols. The medium was replaced two-to-three times every week.
10
Ovarian Cell Line Cultivation
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The normal ovarian cell line IOSE80 was purchased from American Type Culture Collection (ATCC, USA) and cultured in RPMI-1640 medium (HyClone, USA) with 10% fetal bovine serum (FBS, [Biological Industries, Israel]). The human ovarian carcinoma cell lines A2780 and SKOV3 were obtained from ATCC and cultured in DMEM medium and McCoy’s 5A medium (Biological Industries, Israel) containing 10% FBS, respectively. Cells were cultivated at 37°C in a humidified atmosphere containing 5% CO2.
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