HIBEPIC, HCCC-9810, RBE, TFK-1, and HuCC-T1 cells (ATCC, Manassas, VA, USA) were cultured in RPMI-1640 medium supplemented with 10% fetal bovine serum. All cell lines were transfected with plasmids using DharmaFECT Duo Transfection Reagent (Thermo Scientific, Waltham, MA, USA) according to the manufacturer's instructions. Vectors expressing short hairpin RNA (shRNA) sequences were provided by Sangon Biotechnology (Shanghai, China). Cisplatin was obtained from Qilu Pharm (Jinan, China) was obtained from Hansoh (Jiangsu, China).
Tfk 1
Manufactured by American Type Culture Collection
Sourced in United States
The TFK-1 is a laboratory equipment used for cell culture applications. It is designed to facilitate the growth and maintenance of cells in a controlled and sterile environment.
Automatically generated - may contain errors
Lab products found in correlation
Hucct1, by American Type Culture Collection (7 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (6 mentions)
Qbc939, by American Type Culture Collection (5 mentions)
Hccc9810, by American Type Culture Collection (4 mentions)
Huh28, by American Type Culture Collection (4 mentions)
Rpmi 1640 medium, by Thermo Fisher Scientific (4 mentions)
Hibepic, by American Type Culture Collection (2 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (2 mentions)
Cclp1, by American Type Culture Collection (2 mentions)
Dihydromyricetin, by Merck Group (2 mentions)
Dimethyl sulfoxide (dmso), by Merck Group (2 mentions)
Dharmafect duo transfection reagent, by Thermo Fisher Scientific (1 mentions)
Sh eif5a2, by GenePharma (1 mentions)
Dmem medium, by Thermo Fisher Scientific (1 mentions)
Shrna nc, by GenePharma (1 mentions)
Lipofectamine 3000, by Thermo Fisher Scientific (1 mentions)
Lipofectamine 3000 reagent, by Thermo Fisher Scientific (1 mentions)
Si circrna, by GenePharma (1 mentions)
Hucct1, by Thermo Fisher Scientific (1 mentions)
Jurkat, by Thermo Fisher Scientific (1 mentions)
14 protocols using tfk 1
1
Cell Line Transfection and Cisplatin Assay
2
Investigating eIF5A's Role in CCA Cells
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CCA cells (HuCCT1, TFK-1, KKU-452, KKU-100, and QBC939) and human intrahepatic biliary epithelial cells (HIBEpiC) were purchased from ATCC. The DMEM medium (Invitrogen, USA) containing 10% fetal bovine serum (Gibco, USA) was used for cells. The culture conditions were 37°C and 5% CO2 in an incubator.
The cells were inoculated into 6-well plates according to the density of 2.5 × 105/well. sh-eIF5A#1 (5′-GCATTACGTAAGAATGGCTTT-3′), sh-eIF5A#2 (5′-GCATTCAAGATGGTTACCTTT-3′), sh-eIF5A#3 (5′-GCCATGTAAGATCGTCGAGAT-3′), shRNA-NC (5′-TTCTCCGAACGTGTCACGT-3′), pcDNA, and pcDNA-eIF5A were obtained from GenePharma (Shanghai, China). As previous studies [23 (link)], after overnight of culture, the serum-free medium was replaced, and then transfection was carried out by lipofectamine 3000 (Invitrogen). After 6 h of culture, medium was replaced; then the cells were cultured for 24 h. Following that, subsequent experiments were performed.
The cells were inoculated into 6-well plates according to the density of 2.5 × 105/well. sh-eIF5A#1 (5′-GCATTACGTAAGAATGGCTTT-3′), sh-eIF5A#2 (5′-GCATTCAAGATGGTTACCTTT-3′), sh-eIF5A#3 (5′-GCCATGTAAGATCGTCGAGAT-3′), shRNA-NC (5′-TTCTCCGAACGTGTCACGT-3′), pcDNA, and pcDNA-eIF5A were obtained from GenePharma (Shanghai, China). As previous studies [23 (link)], after overnight of culture, the serum-free medium was replaced, and then transfection was carried out by lipofectamine 3000 (Invitrogen). After 6 h of culture, medium was replaced; then the cells were cultured for 24 h. Following that, subsequent experiments were performed.
3
Silencing hsa_circ_0000673 in CCA cells
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The human CCA cell lines RBE, TFK-1, KMBC, and HuH28 were purchased from the American Type Culture Collection (Manassas, VA, USA). Cells were cultured with RPMI-1640 medium (Hyclone, Logan, UT, USA) supplemented with 10% FBS and penicillin/streptomycin at 37° C with 5% CO2.
SiRNA against hsa_circ_0000673 (si-circRNA: 5′-TATTAGACGTTCTTGGTGAAAATCTTTACA-3′) was designed and synthesized by GenePharma (Shanghai, China). Following the manufacturer’s instructions, 50 nM of siRNA was transfected into RBE and KMBC cells using Lipofectamine 3000 reagent (Invitrogen, Carlsbad, CA, USA).
SiRNA against hsa_circ_0000673 (si-circRNA: 5′-TATTAGACGTTCTTGGTGAAAATCTTTACA-3′) was designed and synthesized by GenePharma (Shanghai, China). Following the manufacturer’s instructions, 50 nM of siRNA was transfected into RBE and KMBC cells using Lipofectamine 3000 reagent (Invitrogen, Carlsbad, CA, USA).
4
Cell Line Culture and Validation
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Human embryonic kidney cell line HEK293T, human cholangiocarcinoma cell line TFK‐1 and HuCCT1, and human acute T‐lymphocytic leukemia cell line Jurkat were obtained from American Type Culture Collection (ATCC, Manassas, VA, USA). HEK293T and TFK‐1 cell lines were cultured in Dulbecco's modified Eagle medium (DMEM; GIBCO, ThermoFisher Scientific, Beijing, China) supplemented with 10% fetal bovine serum (FBS; GIBCO) and 1% penicillin‐streptomycin (GIBCO) at 37°C with 5% CO2. HuCCT1 and Jurkat cell lines were cultured in RPMI‐1640 medium (GIBCO) supplemented with 10% FBS and 1% penicillin‐streptomycin at 37°C with 5% CO2. Cell lines were confirmed for surface expression of target antigens and routinely tested for mycoplasma.
5
Culturing Intrahepatic Bile Duct Cells
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Human intrahepatic bile duct epithelial cells (HiBECs) and CCA cell lines (HUCCT1, TFK1, and QBC939) were obtained from American Type Culture Collection (USA). The cells were cultured in Roswell Park Memorial Institute-1640 medium (Gibco, USA) containing 15% fetal bovine serum (Gibco) and 1% penicillin/streptomycin (Gibco) in a humidified incubator containing 5% CO2 at 37°C.
6
Cultivation and Characterization of Cell Lines
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The cell line TFK-1 was cultured in RPMI-1640 supplemented with 10% FBS and antibiotics (penicillin 100 U/ml; streptomycin 100 µg/ml). Mz-ChA-2 cells were cultured in RPMI-1640 supplemented with 10% FBS, 2 mM l -glutamine, 1 × MEM non-essential amino acids (Gibco, Life Technologies, Darmstadt, Germany) and antibiotics (penicillin 100 U/ml; streptomycin 100 µg/ml).
Both cell lines were cultured at 37 °C in a humid atmosphere with 5% CO2 and passaged when 90% confluency was reached (use of Trypsin–EDTA 0.05%, Life Technologies, California, USA). To exclude mycoplasma contamination, both cell lines were routinely checked using PromoKine PCR Mycoplasma Test Kit (Biomedica Medizinprodukte GmbH & Co KG, Vienna, Austria). STR profiling (PowerPlex 16HS System, Promega, Madison USA) was done to verify cell lines. Both cell lines (TFK-1 and Mz-ChA-2 cells) were obtained from the American Type Culture Collection (ATCC).
Both cell lines were cultured at 37 °C in a humid atmosphere with 5% CO2 and passaged when 90% confluency was reached (use of Trypsin–EDTA 0.05%, Life Technologies, California, USA). To exclude mycoplasma contamination, both cell lines were routinely checked using PromoKine PCR Mycoplasma Test Kit (Biomedica Medizinprodukte GmbH & Co KG, Vienna, Austria). STR profiling (PowerPlex 16HS System, Promega, Madison USA) was done to verify cell lines. Both cell lines (TFK-1 and Mz-ChA-2 cells) were obtained from the American Type Culture Collection (ATCC).
7
Cell Culture of Cholangiocarcinoma Lines
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Human normal bile duct cell line and CCA cell lines HuCCT1, RBE, QBC939, TFK‐1, HCCC9810, Huh‐28, and CCLP1 were purchased from American Type Culture Collection (ATCC) and were cultured in RPMI‐1640 medium (Pricella Life Science & Technology Co., Ltd) supplemented with 10% fetal bovine serum (Scitecher, Beijing Dingguo Changsheng Biotechnology Co., Ltd) at 37°C in a 5% CO2 incubator.
8
Regulation of Lnc-ATB and miR-200c in CCA
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Human bronchial epithelial cell BEC and four CCA cell lines HUCCT1, RBE, TFK1 and Huh-28 were obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA) and cultured in DMEM supplement with 10% FBS (GIBCO, Waltham, MA, USA) and 1% penicillin/streptomycin. All the cells were cultured at 37°C water-saturated 5% CO2 atmosphere.
The oligonucleotide sequences of miR-200 mimics, inhibitor or negative control were purchased from GenePharma (Shanghai, China). siRNA-Lnc-ATB or negative control were conducted by RiboBio (Guangzhou, China). psiCHECK2-UTR (wild-type or mutant miR-200c target site) of Lnc-ATB was conducted by GenePharma. The lentivirus vector of siRNA-Lnc-ATB, miR-200c inhibitor or negative control was conducted by GeneChem (Shanghai, China). The antibody to CCND1 (#ab16663), CDK2 (#ab32147), and PCNA (#ab92552) was obtained from Abcam (Massachusetts, USA), and caspase-3 (#9664), BCL-2 (#9662), ZEB1/2, E-cadherin and N-cadherin (EMT antibody sampler Kit #9782) were obtained from Cell Signaling Technology (Denver, MA, USA).
The oligonucleotide sequences of miR-200 mimics, inhibitor or negative control were purchased from GenePharma (Shanghai, China). siRNA-Lnc-ATB or negative control were conducted by RiboBio (Guangzhou, China). psiCHECK2-UTR (wild-type or mutant miR-200c target site) of Lnc-ATB was conducted by GenePharma. The lentivirus vector of siRNA-Lnc-ATB, miR-200c inhibitor or negative control was conducted by GeneChem (Shanghai, China). The antibody to CCND1 (#ab16663), CDK2 (#ab32147), and PCNA (#ab92552) was obtained from Abcam (Massachusetts, USA), and caspase-3 (#9664), BCL-2 (#9662), ZEB1/2, E-cadherin and N-cadherin (EMT antibody sampler Kit #9782) were obtained from Cell Signaling Technology (Denver, MA, USA).
9
Prodigiosin Modulates CCA Cell Lines
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Human CCA cancer cell lines (TFK-1, HuCCT-1, HUH28, QBC939, RBE, and CCLP-1) were purchased from the American Type Culture Collection (ATCC; Manassas, VA, USA) and cultured in RPMI1640 medium supplemented with 10% fetal bovine serum (FBS, Biological Industries, Israel) at 37 °C. STX17 overexpression (STX17), SNAP29 overexpression (SNAP29) and negative-control (Vector) lentiviruses were purchased from Genechem (Shanghai, China). All transfections were performed base on the instructions of manufacturer. All cell lines have been tested for mycoplasma contamination before conducting this study. Prodigiosin was purchased from Sigma-aldrich (CAS: 56144-17-3, USA) and stored at –20 °C.
10
Cell line authentication and maintenance
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Human embryonic kidney HEK293T cells (CVCL_3216), human hepatocellular carcinoma HepG2 cells (CVCL_0027), human CCA KKU‐100 (CVCL_3996), KKU‐213 (CVCL_M261), SNU‐308 (CVCL_5048), SNU‐1079 (CVCL_5008), SNU‐1196 (CVCL_5015), TFK‐1 (CVCL_2214) and TKKK (CVCL_5599) cells were obtained from American Type Culture Collection (ATCC, Manasssa, VA, USA), Korea Cell Line Bank (KCLB, Seoul, Korea) or Japanese Cancer Research Resources Bank (JCRB, Osaka, Japan). The cell lines were maintained in RPMI‐1640 medium or DMEM (WELGENE, Daegu, Korea) supplemented with 10% fetal bovine serum (GIBCO, Grand Island, NY, USA) and 1% penicillin–streptomycin (WELGENE), in a 5% CO2 incubator, at 37°C. All cell lines were tested for the presence of mycoplasma using the EZ‐PCR™ Mycoplasma Detection Kit (Biological Industries, #20–700‐20, Israel). Cell line authentication was performed via short tandem repeat (STR) profiling by KCLB.
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