293T, HepG2, Hep3B and HuH7 were purchased from Shanghai Institute of Biochemistry and Cell Biology. Hep3B cells were maintained in Minimum Essential Medium (cat. no. 41500034; HyClone) supplemented with 10% FBS (cat. no. P30-3302, PAN-Biotech). HepG2 and HuH7 cells were maintained in DMEM (cat. no. C0006; Hangzhou Keyi Biotechnology Co., Ltd.) supplemented with 10% FBS. 293T cells were cultured in 90% DMEM supplemented with 10% FBS. The hypoxic condition was created by placing liver cancer cells in a sealed hypoxic chamber and equilibrating it with a gas mixture of 1% O2, 5% CO2 and 94% N2. To prevent mycoplasma contamination, mycoplasma testing was performed for the cell lines used. Mycoplasma testing was performed on cell lines, which were then authenticated for genotypes using short tandem repeat DNA fingerprinting; the cells were passaged for <6 months (17 (link)).
Hep3b
Manufactured by Shanghai Institute of Biochemistry and Cell Biology
Sourced in China
Hep3B is a cell line derived from a human hepatocellular carcinoma. It is a widely used in vitro model for the study of liver cancer and related biological processes.
Automatically generated - may contain errors
Lab products found in correlation
Huh 7, by Shanghai Institute of Biochemistry and Cell Biology (10 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (10 mentions)
Hepg2, by Shanghai Institute of Biochemistry and Cell Biology (8 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (6 mentions)
Hcclm3, by Shanghai Institute of Biochemistry and Cell Biology (5 mentions)
Mhcc 97h, by Shanghai Institute of Biochemistry and Cell Biology (4 mentions)
Hl 7702, by Shanghai Institute of Biochemistry and Cell Biology (4 mentions)
Dmem rpmi1640, by Thermo Fisher Scientific (3 mentions)
Penicillin streptomycin mixture, by Thermo Fisher Scientific (3 mentions)
Bel 7402, by Shanghai Institute of Biochemistry and Cell Biology (3 mentions)
Bel 7404, by Shanghai Institute of Biochemistry and Cell Biology (3 mentions)
Smcc7721, by Shanghai Institute of Biochemistry and Cell Biology (3 mentions)
Snu182, by Shanghai Institute of Biochemistry and Cell Biology (2 mentions)
L glutamine, by Thermo Fisher Scientific (2 mentions)
Puromycin, by Merck Group (1 mentions)
Penicillin streptomycin, by Wisent (1 mentions)
Penicillin, by Tiangen Biotech (1 mentions)
Streptomycin, by Tiangen Biotech (1 mentions)
Snu449, by Shanghai Institute of Biochemistry and Cell Biology (1 mentions)
Lipofectamine 2000, by Thermo Fisher Scientific (1 mentions)
17 protocols using hep3b
1
Establishing Hypoxic Liver Cancer Cell Lines
2
Genetic Manipulation of Hep3B Cells
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Human HCC cell line (Hep3B) was purchased from the Shanghai Institute of Biochemistry and Cell Biology and cultured in Dulbecco’s modified Eagle’s medium (DMEM) with 10% fetal bovine serum (FBS) and 1% penicillin-streptomycin (WISENT, Shanghai, China). Before the cells were harvested, they were cultured at 37 °C with 5% CO2. The Cas9 expression construct pLV1-CMV-Cas9-Puro-U6-sgRNA was used for the expression of sgRNAs in Hep3B cells; sgRNA targeting TBC1D8 was 5′-AAATGGAGCGACCCGTGCAT-3′, and sgRNA targeting TBC1D14 was 5′-GGAATCCCTCCAAGTGTGAG-3′. Empty pLV1-CMV-Cas9-Puro-U6-sgRNA plasmid was used as a control (sgNC). Puromycin (1 μg/ml, Merck, 540,411) was added 24 h after transfection. Cells were collected 72 h after transfection for subsequent experiments.
3
HCC Cell Lines and Tissue Specimens
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Five HCC cell lines (HepG2, Hep3B, SNU182, SNU449, MHCC-97H) and CL-48 normal hepatocytes all were purchased from Shanghai Institute of Cell Biology (Shanghai, China) and cultured in RMPI-1640 medium with 3% fetal bovine serum (FBS; Tiangen, Shanghai, China) plus 100 U/ml penicillin and streptomycin (100 µg/ml) purchased from Tiangen. The surgically sectioned HCC specimens were acquired from The First Affiliated Hospital of Wenzhou Medical University (Wenzhou, China) from July 2010 to October 2014. Immediately after surgical resection, these tissues were stored at −80°C until usage. None patients received preoperative chemotherapy or radiotherapy. All patients have signed formal consent forms. The research for human samples was approved by Ethics Committee of The First Affiliated Hospital of Wenzhou Medical University.
4
Modulation of Hepatocellular Carcinoma by CircFOXM1 and SPAG5
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Human HCC cell lines (Huh7, HepG2, Hep3B, HCCLM3 and MHCC97-H) and normal human hepatocytes (MIHA) were obtained from Shanghai Institute of Cell Biology (Shanghai, China). Cells were maintained in DMEM or RPMI-1640 medium (HyClone) containing 10% fetal bovine serum (FBS) under the condition of 5% CO2 and 37 °C.
The specific circFOXM1 or SPAG5 siRNA was designed and synthesized by GenePharma (Shanghai, China). The sequence of circFOXM1 or SPAG5 was inserted in pLCDH-ciR to construct the recombinant overexpression vector respectively. HCC cells were transfected with circFOXM1 or SPAG5 siRNA, pLCDH-circFOXM1 or pLCDH-SPAG5, miR-1179 mimics or miR-1179 inhibitor using Lipofectamine 2000 (Invitrogen), following the manufacturer’s instructions.
The specific circFOXM1 or SPAG5 siRNA was designed and synthesized by GenePharma (Shanghai, China). The sequence of circFOXM1 or SPAG5 was inserted in pLCDH-ciR to construct the recombinant overexpression vector respectively. HCC cells were transfected with circFOXM1 or SPAG5 siRNA, pLCDH-circFOXM1 or pLCDH-SPAG5, miR-1179 mimics or miR-1179 inhibitor using Lipofectamine 2000 (Invitrogen), following the manufacturer’s instructions.
5
Culturing Human Liver Cell Lines
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The human normal hepatocyte cell lines L‐02 and the HCC cell lines Hep3B and Huh‐7 were purchased from the Shanghai Institute of Cell Biology at the Chinese Academy of Sciences (Shanghai, China). The HCC cell lines SMMC‐7721, Bel‐7402, Bel‐7404, SK‐Hep‐1, and HepG2 were obtained from Genechem Co., Ltd. (Shanghai, China). The HCC cell line MHCC‐97L was obtained as a gift from the First Affiliated Hospital of Xi'an Jiaotong University. MHCC‐97L, Huh‐7, and HepG2 cells were cultured in DMEM containing 10% FBS. L‐02, SMMC‐7721, Bel‐7402, and Bel‐7404 cells were cultured in RPMI‐1640 medium, while Hep3B and SK‐Hep‐1 cells were cultured in MEM medium containing 10% FBS. All media were supplemented with penicillin (100 U/ml) and streptomycin (100 U/ml). All cells were maintained in an incubator with a humidified atmosphere of 5% CO2 at 37°C.
6
Authenticating Human HCC Cell Lines
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Human HCC cell lines (Hep3B, Huh7 and PLC) were purchased from Shanghai Institute of Cell Biology, Chinese Academy of Sciences. The authenticity of all the cell lines was verified by short-tandem repeat profiling (Invitrogen, CA, USA), and the cell lines were tested for Mycoplasma contamination (MycoAlert, Lonza, Basel, Switzerland). All cell lines were cultured at 37 ℃ in an atmosphere containing 5% CO2 in MEM or DMEM (Gibco, NY, USA) supplemented with 10% fetal bovine serum (Wisent, Montreal, Canada).
7
Acquisition of HCC and Adjacent Tissues
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HCC and adjacent normal tissues were acquired from 45 patients, 33 males and 12 females, who had not received therapy prior to surgery during the period from 2018 to 2020 at the affiliated hospital of Nantong University. The study was conducted in accordance with the Declaration of Helsinki (as revised in 2013). Informed consent was obtained from the patients, and this research was approved by the Ethics Committee of Affiliated Hospital of Nantong University (ethical code: 2018-L006). Samples were frozen in liquid nitrogen immediately and stored at –80 °C for future use. Patients provided written informed consent prior to surgery. HCC cell lines (Hep3B, Huh-7, Li-7, and SUN182) were purchased from Shanghai Institute of Cell Biology (Shanghai, China) and a normal liver cell line (QSG-7701) was purchased from Beyotime (Shanghai, China). Cells were cultured at 37 °C under 5% CO2 in RPMI-1640 or DMEM (Thermo Fisher Scientific, Waltham, MA, USA) containing 10% fetal bovine serum (FBS; Thermo Fisher Scientific), streptomycin (100 µg/mL) and penicillin (100 U/mL).
8
Authenticated Human Hepatocellular Carcinoma Cell Lines
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Human HCC cell lines (BEL-7402, BEL-7404, Huh7, Hep3B, and HepG2) and the normal hepatocyte cell line HL-7702 were purchased from the Shanghai Institute of Cell Biology (Shanghai, China) with a certificate of authenticity for each cell line. The samples were maintained in DMEM/RPMI 1640 (Gibco BRL, Grand Island, NY, United States) with 10% heat-inactivated fetal calf serum, 2 mM l -glutamine, and 100 U/mL penicillin–streptomycin mixture (Gibco BRL, Grand Island, NY, United States) at 37°C in 5% CO2. After the cells were cultured for several months, all cell lines were evaluated for genetic identification prior to the experiments. Based on the appraisal reports provided by CoBioer Biosciences Co., Ltd. (Nanjing, China), no mutations or contaminations were found in our cell lines.
9
Cell Culture of Human Liver Lines
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Human HCC cell lines (Huh7, HCCLM3, SMCC7721 and Hep3B) were purchased from the Shanghai Institute of Cell Biology, China. The immortalized liver cell line HL-7702 was purchased from Shanghai Fu Xiang Biotechnology Co., Ltd. The cells were cultured in DMEM (Gibco) supplemented with FBS (Hyclone) to a final concentration of 10% and were exposed to antibiotics at 37 °C with 5% CO2.
10
Authenticity Evaluation of Hepatocyte Cell Lines
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The human HCC cell lines (BEL-7402, BEL-7404, Huh7, Hep3B and HepG2) and normal hepatocyte cell HL-7702 were purchased from the Shanghai Institute of Cell Biology with a certificate of authenticity for each cell line. The samples were maintained in DMEM/RPMI1640 (Gibco BRL, Grand Island, NY, USA) supplemented with 10 % heat-inactivated fetal calf serum, 2 mM L-glutamine, and 100 U/mL penicillinstreptomycin mixture (Gibco BRL, Grand Island, NY, USA) at 37 ℃ in 5 % CO2. After undergoing cell culture in our laboratory for several months, all cell lines used were evaluated for genetic identification prior to the experiments. Based on the appraisal reports provided by CoBioer Biosciences Co., Ltd., no mutations and contaminations were found in our cell lines.
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