The human colon cancer cell lines KM12, HT29, HCT116, SW480, and HCT15 were obtained from the National Cancer Institute (NCI, Frederick, MD, USA). The fibrosarcoma cell line HT1080 was purchased from Korean Cell Line Bank (KCLB, Seoul, Korea), and HEK293T was obtained from American Type Culture Collection (ATCC, Manassas, VA, USA). TRI‐DR U2OS was a gift from Dr. Oberdoerffer [21 (link)]. The cells were cultured in DMEM or RPMI 1640 supplemented with 10% FBS (Thermo Fisher Scientific, Waltham, MA, USA) in cell culture‐treated flasks for maintenance and for BCC culture. CSC was enriched in DMEM/F‐12 (Thermo Fisher Scientific) supplemented with EGF, basic fibroblast growth factor, and B27 supplement on poly HEMA (polymer of 2‐hydroxyethyl methacrylate)‐treated culture dishes as previously described [22 (link)]. Both media were supplemented with 1% penicillin and streptomycin (Welgene, Gyeongsan, Korea).
Ht1080
Manufactured by Korean Cell Line Bank
Sourced in United States
The HT1080 is a cell line derived from a human fibrosarcoma. It is a widely used in vitro model for the study of cancer biology and drug screening. The HT1080 cell line exhibits rapid growth and has been characterized for various cellular and molecular properties.
Automatically generated - may contain errors
Lab products found in correlation
Mcf 7, by Korean Cell Line Bank (3 mentions)
Mda mb 231, by Korean Cell Line Bank (3 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (3 mentions)
Hep3b, by Korean Cell Line Bank (2 mentions)
Skov3, by Korean Cell Line Bank (2 mentions)
Polybrene, by Merck Group (2 mentions)
Dmem f12, by Thermo Fisher Scientific (2 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (2 mentions)
Streptomycin, by Welgene (1 mentions)
Rpmi 1640, by Thermo Fisher Scientific (1 mentions)
Hek293t, by American Type Culture Collection (1 mentions)
Penicillin, by Welgene (1 mentions)
Penicillin, by Thermo Fisher Scientific (1 mentions)
Streptomycin, by Thermo Fisher Scientific (1 mentions)
Complete dmem, by Thermo Fisher Scientific (1 mentions)
Du145, by Korean Cell Line Bank (1 mentions)
Hepg2, by Korean Cell Line Bank (1 mentions)
Lipofectamine, by Thermo Fisher Scientific (1 mentions)
Aspc 1, by Korean Cell Line Bank (1 mentions)
Hct116, by Korean Cell Line Bank (1 mentions)
10 protocols using ht1080
1
Culturing Colon Cancer and Fibrosarcoma Cells
2
Long-term Culture of Fibrosarcoma Cells
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Human fibrosarcoma cell line HT1080 was obtained from the Korean Cell Line Bank, and the human embryonic kidney cell line 293FT was obtained from Invitrogen (Thermo Fisher Scientific, USA). All cells were grown in Dulbecco’s modified Eagle’s medium (DMEM), supplemented with 10% fetal bovine serum (FBS) and 1%streptomycin/penicillin, and cultured in a humidified 5% CO2 incubator at 37°C. For the cell viability assay, transduced HT1080 cells were sub-cultured at 1:10 or 1:8 dilution every 3 to 4 days for up to 28 days.
3
Cell Culture Protocol for Cancer Cell Lines
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HT1080, HeLa, MCF7, HepG2, MDA-MB-231, and DU145 cells purchased from the Korean Cell Line Bank (Seoul, Korea). The cells were cultured in DMEM complete (Gibco BRL, Grand Island, NY, USA) with 10% fetal bovine serum (FBS; Gibco BRL), 100 U/mL penicillin, and 100 μg/mL streptomycin (Invitrogen, Waltham, MA, USA). The cells were cultured at 37 °C in a humidified atmosphere with 5% CO2.
4
Cell Culture and Fluorescent Labeling
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The HepG2 (human hepatocellular carcinoma), Hep3B (human hepatocellular carcinoma), SK-OV-3 (human ovarian cancer), HT-1080 (human fibrosarcoma), HFF (human fibroblast), MDA-MB-231(human triple-negative breast cancer), and MCF7 (human luminal type breast cancer) cell lines were purchased from the Korean Cell Line Bank. All cell lines were cultured in Dulbecco’s modified Eagle’s medium (DMEM; Gibco, USA) containing 10% fetal bovine serum (FBS; Gibco), 10 U/mL penicillin, and 10 μg/mL streptomycin in a humidified atmosphere with 5% CO2 at 37°C. EV-free FBS, from which EVs were removed by ultracentrifugation, was used in all experiments related to the coculture and isolation of EVs. MDA-MB-231 cells were transfected with a lentiviral vector encoding the red fluorescent protein palmitoylated tandem dimer Tomato (tdTomato) using Lipofectamine (Life Technologies). The supernatant was recovered at 48 h after transfection, filtered through a 0.45-μm membrane, and then added to plated MDA-MB-231 cells supplemented with 4 μg/mL Polybrene (Sigma–Aldrich). After viral infection, tdTomato-positive cells were selected using fluorescence-activated cell sorting for stable integration of the transfected construct.
5
Cell Line Cultivation and Sourcing
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The Hep3B, SKOV-3, U-118MG, U-87MG, FaDu, MCF-7, T-47D, DU145, A549, B16F10, LLC1, CT26.WT, HeLa, and NHBE (normal human bronchial epithelial) cell lines were purchased from the American Type Culture Collection (ATCC, VA). The HCT116, SW620, HT-1080, AsPC-1, MIA PaCa-2, and JC cell lines were purchased from the Korea Cell Line Bank (Seoul, Korea). A2780 and Vero cells were purchased from the European Collection of Authenticated Cell Cultures (ECACC, Salisbury, United Kingdom) and the Korean Ministry of Food and Drug Safety (Chungcheongbuk-do, Korea), respectively. All cells were cultured in an incubator at 37°C with 5% CO2 in distinct culture media (Supplementary Table 1 ) supplemented with fetal bovine serum (FBS) (SH30084.03; GE Healthcare Life Sciences, PA), antibiotic/antimycotic (A/A; 15240112), l -glutamine (25030164), and human insulin (12585014) (all from Thermo Fisher Scientific, MA).
6
HT1080 Fibrosarcoma Cell Viability Assay
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Human fibrosarcoma cell line, HT1080, was adopted from KCLB (Korea Cell Line Bank, Jongro, Seoul, Korea), and cultured in T-75 flasks (SPL, Pocheon, Gyeonggi, Korea) in 5% CO2 and at 37 °C in a humidified incubating condition, and the medium used was RPMI 1640 (GenDEPOT, Baker, TX, USA) with 10% fetal bovine serum (FBS) (GenDEPOT, Baker, TX, USA) and 100 unit/mL penicillin–streptomycin (Gibco-BRL, Grand Island, NY, USA). The cell lines were washed with PBS buffer (Gibco-BRL, Grand Island, NY, USA) and the medium was changed six times a week.
For cell viability assay, cell lines were transferred to 96-well plates at 5 × 103 cells/well density. After their transfer, cells were cultured for 24 h and their medium was changed with fresh medium, which was followed by the treatment with samples (100, 50 and 10 µM). Cells were re-supplied with fresh medium after 24 h of incubation and treated with 100 µL of MTT solution (1 mg/mL), and further incubated for 4 h. Finally, the wells were aspirated and introduced into 100 µL of dimethyl sulfoxide (DMSO), in order to solubilize the formazan crystals for the measurement by a Victor3 reader (PerkinElmer, Waltham, MA, USA) at 540 nm optical density.
For cell viability assay, cell lines were transferred to 96-well plates at 5 × 103 cells/well density. After their transfer, cells were cultured for 24 h and their medium was changed with fresh medium, which was followed by the treatment with samples (100, 50 and 10 µM). Cells were re-supplied with fresh medium after 24 h of incubation and treated with 100 µL of MTT solution (1 mg/mL), and further incubated for 4 h. Finally, the wells were aspirated and introduced into 100 µL of dimethyl sulfoxide (DMSO), in order to solubilize the formazan crystals for the measurement by a Victor3 reader (PerkinElmer, Waltham, MA, USA) at 540 nm optical density.
7
Cell Culture Conditions for Fibrosarcoma and Adenocarcinoma
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Human fibrosarcoma (HT-1080) and colorectal adenocarcinoma cells (HT-29) were obtained from the Korean Cell Line Bank (Seoul, Korea). Typically, the cells were cultured in T-25 tissue culture flasks (SPL, Korea) in RPMI-1640 media containing 10% FBS and 1 × P/S and incubated at 37 °C in a humidified incubator with a 5% CO2 atmosphere. When the cells reached 80% confluence, the culture media were changed to fresh serum-free conditioned media. For immunoblotting and zymography, the media were collected 48 h after culture and concentrated by using an Amicon ultra centrifugal filter unit (50 kDa MWCO, Millipore, Billerica, MA, USA) and by centrifugation (1500× g for 20 min).
8
Cytotoxicity Evaluation of Synthesized Compounds
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Human fibroblast HT1080 cells were procured from the Korean Cell Line Bank in Seoul, Korea, and utilized for evaluating the cytotoxicity of the synthesized compounds (5a-k). The cultivation and treatment procedures were performed according to the methods described in the literature [52] .
9
Establishing Fluorescent Cancer Cell Lines
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HepG2 (human hepatocellular carcinoma), Hep3B (human hepatocellular carcinoma), SK-OV-3 (human ovarian cancer cell), HT-1080 (human fibrosarcoma cell), HFF (human fibroblast), MDA-MB-231(human triple negative breast cancer cell) and MCF7 cells (human luminal type breast cancer cell) were purchased from Korean Cell Line Bank. The cells were cultured in Dulbecco's modified Eagle's medium (DMEM) containing with 10% fetal bovine serum (FBS), 10 U/ml penicillin, and 10 μg/ml streptomycin under humidified 5% CO 2 atmosphere and at 37 °C.
MDA-MB-231 were transfected with a lentiviral vector encoding the red fluorescent protein palmitoylated-tdTomato (tdTomato) using lipofectamine transfection reagent (Life Technologies). Supernatant recovered after 48h from 293T transfected cells was filtered by a 0.45 μm pore membrane and added to MDA-MB-231 plated cells supplemented with 4 μg/mL polybrene (Sigma-Aldrich). After viral infection, palm-td-Tomato positive cells were selected using FACS for stable integration of the transfects.
MDA-MB-231 were transfected with a lentiviral vector encoding the red fluorescent protein palmitoylated-tdTomato (tdTomato) using lipofectamine transfection reagent (Life Technologies). Supernatant recovered after 48h from 293T transfected cells was filtered by a 0.45 μm pore membrane and added to MDA-MB-231 plated cells supplemented with 4 μg/mL polybrene (Sigma-Aldrich). After viral infection, palm-td-Tomato positive cells were selected using FACS for stable integration of the transfects.
10
Cultivation and Maintenance of Human and Murine Cell Lines
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Cells and animals. Human fibrosarcoma HT1080 (KCLB no. 10121) and murine melanoma B16F10 (KCLB no. 80008) cells were purchased from the Korean Cell Line Bank (Seoul, Korea) and maintained in RPMI-1640 or Dulbecco's modified eagle's medium (DMeM; Cellgro, Manassas, VA, uSA) supplemented with 10% (v/v) fetal bovine serum (FBS; Cellgro), 100 u/ml penicillin and 100 µg/ml streptomycin (Cellgro) at 37˚C in a humidified incubator with 5% CO 2 . Human umbilical vein endothelial cells (HuVeCs) were purchased from InnoPharmaScreen (Asan, Korea) at passage 2, maintained in endothelial cell growth medium-2 (egM-2; PromoCell, Heidelberg, germany), and used for experiments at passages 3-8. For the animal experiments, female athymic nude mice were purchased from nara Biotech (Seoul, Korea). Female C57BL/6 mice and male Sprague Dawley rats were obtained from Taconic Farms Inc. (Samtako Bio Korea, Osan, Korea). All animals were housed under controlled conditions using a 12/12-h light/dark cycle at 22±1˚C and 55±5% humidity under specific pathogen-free conditions. All animal experiments were approved by the Animal Care and use Committee of the Korea Institute of Oriental Medicine (KIOM, Daejeon, Korea), with the reference numbers #13-42, #13-48 and #14-27, and were performed in accordance with the guidelines of the Animal Care and use Committee at KIOM.
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