Hct 8

Manufactured by Keygen Biotech

Sourced in China

The HCT-8 is a laboratory instrument designed for the measurement and analysis of cell samples. It is capable of performing high-content screening and provides quantitative data on various cellular parameters, including cell count, viability, and morphology. The HCT-8 is a versatile tool that can be used in a wide range of applications, such as drug discovery, toxicology studies, and basic cell biology research.

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HCT-8/5-FU cells (3 citations) HCT-8 cells (2 citations)

20 protocols using hct 8

Establishment and Characterization of Cancer Cell Lines

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All cell lines used were of human origin. A549 (non-small cell lung cancer) and MCF-7 (breast cancer) cell lines were purchased from Cell Resource Center of Shanghai Institute for Biological Sciences (Chinese Academy of Sciences, Shanghai, China). A549T (Taxol-resistant A549 subline) and MCF-7R (Adriamycin-resistant MCF-7 subline) cell lines were from Shanghai Aiyan Biological Technology Co. Ltd (Shanghai, China). HCT-8 (colon carcinoma), A2780 (ovarian cancer), HCT-8T (Taxol-resistant HCT-8 subline) and A2780T (Taxol-resistant A2780 subline) were from Nanjing KeyGEN BioTECH Co. Ltd (Nanjing, China). PC-3 (prostate cancer) was from Typical Culture Preservation Commission Cell Bank, Chinese Academy of Sciences (Shanghai, China).
All cell lines were routinely cultured at 37 °C in humidified atmosphere with 5% CO₂. Unless stated otherwise, the growth medium used for A549, A549T, MCF-7R, HCT-8, HCT-8T, PC-3 and A2780T cells was RPMI-1640 (Corning) containing 10% fetal bovine serum (FBS, Gibco) and 1% penicillin/streptomycin (Corning). MCF-7 and A2780 cells were propagated in Dulbecco’s Modified Eagle Medium (DMEM, Corning) with 10% fetal bovine serum (FBS, Gibco) and 1% penicillin/streptomycin (Corning). For comparison, some experiments replaced FBS with other serums (calf serum, bovine serum, goat serum, horse serum or human serum) and the same concentration.

Wang L., Liu Y., Qi C., Shen L., Wang J., Liu X., Zhang N., Bing T, & Shangguan D. (2018). Oxidative degradation of polyamines by serum supplement causes cytotoxicity on cultured cells. Scientific Reports, 8, 10384.

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Cell Line Establishment Protocol

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Cell lines used in this study, which included lines derived from hepatocellular carcinoma cells (HpG2), lung cancer cells (NCI-A549, hereafter referred to as A549 cells; NCI-H1299, hereafter referred to as H1299 cells), breast cancer cells (MCF-7), colorectal cancer cells (HCT-8), and pancreatic cancer cells (PANC-1), were obtained from KeyGen Biotech (Nanjing, China). All cells were cultured in RPMI 1640 or DMEM media supplemented with 10% fetal bovine serum under conditions of 37°C, 95% humidity, and 5% CO₂ (hereafter referred to as standard cell culture conditions). No primary human tumor specimens were used in this study.

Yang L., Jin W.Q., Tang X.L., Zhang S., Ma R., Zhao D.Q, & Sun L.W. (2022). Ginseng-derived nanoparticles inhibit lung cancer cell epithelial mesenchymal transition by repressing pentose phosphate pathway activity. Frontiers in Oncology, 12, 942020.

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Culturing of Colorectal Cell Lines

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The human CRC cell lines (HCT116, SW620, SW480, HCT8, and LoVo) and normal colonic epithelial cell line (FHC) were obtained from the Chinese Academy of Science Cell Bank (Shanghai, China) and the FHC human normal colorectal epithelial cell line was sourced from the American Type Culture Collection (ATCC, Manassas, VA, USA). All cell lines were cultured in the appropriate basal medium supplemented with FBS (Gibco, Waltham, MA, USA) and 1% antibiotic/antimycotic solution, then incubated at 37 °C with 5% CO₂ in a humidified atmosphere. FHC and HCT116 cells were cultured in McCoy's 5A medium (KeyGEN, Nanjing, China), LoVo, and HCT8 cells in DMEM, and SW620 and SW480 cells in L-15 media (KeyGEN, Nanjing, China).

Liu J., Zhang X., Yang M, & Zhang X. (2024). CircCOL1A1 promotes proliferation, migration, and invasion of colorectal cancer (CRC) cells and glutamine metabolism through GLS1 up-regulation by sponging miR-214-3p. Journal of Cancer Research and Clinical Oncology, 150(4), 211.

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Characterizing Colorectal Cancer Cell Lines

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The human CRC cell lines SW620, SW480, HCT-8, 5-FU-resistant HCT-8 cells (HCT-8/5-FU) and LoVo were obtained from KeygenBiotech Co. Ltd. (Nanjing, China), which was recently authenticated as truly CRC cell lines. SW620 and SW480 were cultured in Leibovitz’s L-15 (Gibco, Grand Island, NY) medium contained 10% in activated fetal bovine serum (Gibco, Grand Island, NY). 5-FU was added to LoVo cell culture in stepwise increasing concentrations for over 6 months, namely LoVo/5-FU. HCT-8/5-FU and LoVo/5-FU were maintained in RPMI1640 supplementd with 114.7 μM and 107.0 μM 5-FU, respectively. CRC cells were incubated at 37 °C containing 5%CO₂. All cells lines were routinely tested for mycoplasma, which were shown to be negative.

Liu B., Pan S., Xiao Y., Liu Q., Xu J, & Jia L. (2018). LINC01296/miR-26a/GALNT3 axis contributes to colorectal cancer progression by regulating O-glycosylated MUC1 via PI3K/AKT pathway. Journal of Experimental & Clinical Cancer Research : CR, 37, 316.

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Culturing Human Colon Cell Lines

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The human normal colonic epithelial cell line (FHC) was purchased from the American Type Culture Collection (ATCC; Manassas, VA, USA). Human CRC cell lines (HT29, RKO, HCT116, SW480, SW620, LoVo, HCT8 and LS123) were obtained from KeyGEN BioTECH (Nanjing, Jiangsu, China). The cell lines were cultured in appropriate medium supplemented with 10% fetal bovine serum (FBS; Biological Industries, Israel) and 1% antibiotics (100 U/mL penicillin and 100 mg/mL streptomycin; Life Technologies, Inc., Grand Island, NY, USA). All cell lines were maintained in an incubator at 37°C in a humidified atmosphere with 5% CO₂. All cell experiments were performed using mycoplasma-free cells.

Long F., Li L., Xie C., Ma M., Wu Z., Lu Z., Liu B., Yang M., Zhang F., Ning Z., Zhong C., Yu B., Liu S., Wan L., Tian B., Yang K., Guo Y., Chen M., Chou J., Li X., Hu G., Lin C, & Zhang Y. (2023). Intergenic CircRNA Circ_0007379 Inhibits Colorectal Cancer Progression by Modulating miR-320a Biogenesis in a KSRP-Dependent Manner. International Journal of Biological Sciences, 19(12), 3781-3803.

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Colorectal Cancer Tissue Analysis

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Human primary CRC and adjacent tissues were extracted from 36 patients who underwent surgical resections from the First A liated Hospital of Dalian Medical University. All tumor tissues were con rmed by histological examination as adenocarcinoma of the colorectal. Tissues were stored in liquid nitrogen for Quantitative Real-Time PCR (QRT-PCR) and immunohistochemistry experiments. The research protocol was approved by the Ethics Committee of the First A liated Hospital of Dalian Medical University (YJ-KY-FB-2016-16). CRC cell lines HCT-8, Caco2, SW480 SW620, LOVO and human normal colorectal epithelial cell line (FHC) were obtained from KeyGEN Company (Nanjing, Jiangsu, China) and maintained as previously described [21] .

Qi Y., Shan Y., Li S., Huang Y., Guo Y., Huang T., Zhao X, & Jia L. (2022). LncRNA LEF1-AS1/LEF1/FUT8 Axis Mediates Colorectal Cancer Progression by Regulating α1, 6-Fucosylationvia Wnt/β-Catenin Pathway. Digestive diseases and sciences, 67(6).

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Comprehensive Chemical Analysis of Anticancer Compounds

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All commercially available chemicals were from Sigma Chemical Co. and purified when necessary. The purity of all compounds was identified with TLC (Qingdao silica gel F254, 0.25 mm layer thickness) or HPLC (Waters, C18 column, 4.6×150 mm). Bruker Avance II-300 or Bruker Avance III-800 spectrometer was used to record the ¹H NMR (300 or 800 MHz) and ¹³C NMR (75or 200 MHz) spectra, while DMSO-d₆ was the solvent and tetramethylsilane was the internal standard. ZQ 2000 mass spectrometer (Waters, US) or Fourier transform ion cyclotron resonance (FT-ICR, 9.4T solariX, Bruker, US) with dual ion source of ESI/matrix-assisted laser desorption ionization ESI/MS was used for mass analyses.
HCT-116, LS174T, SW620, SGC7901, Eca109, MKN28, HCT-8, and S180 cells were purchased from key GENBioTECH (Nanjing China). Male ICR mice (22 ± 2 g) were purchased commercially from Laboratory Animal Center of Capital Medical University. In vitro and in vivo assays were examined by the Ethics Committee of Capital Medical University. The committee approved that the assays can use the mentioned cells and mice can be used for the assays, and assured that the welfare of mice met the requirements of Animal Welfare Act and NIH Guide for Care and Use of Laboratory Animals.
Biological data were statistically analyzed with ANOVA, and the P-value less than 0.05 was considered statistically significant.

Li G., Ren Y., Zhang X., Zhao S., Wang Y., Wu J., Peng S, & Zhao M. (2019). 13-[CH2CO-Cys-(Bzl)-OBzl]-Berberine: Exploring The Correlation Of Anti-Tumor Efficacy With ROS And Apoptosis Protein. OncoTargets and therapy, 12, 10651-10662.

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Overexpression of COMP in Cancer Cells

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HCT116, HCT-8, and SW620 cells were purchased from KeyGEN BioTECH. Cells were cultured in RPMI 1640 or DMEM containing 10% FBS (Gibco, Life Technologies) and 1% penicillin and streptomycin (Hycult, Life Technologies). The cells were cultured in a cell incubator at 37 ℃ with 5% CO₂, digested by trypsin, and subcultured every 2 days. Full-length COMP or mock (empty vector) or shCOMP was stably expressed in both cell lines by transfection with Lipofectamine 2000 (Invitrogen, USA) and selection with hygromycin or puromycin (Invitrogen, USA). COMP expression was verified by WB. Single clones with good COMP expression were chosen for further experiments.

Zhong W., Hou H., Liu T., Su S., Xi X., Liao Y., Xie R., Jin G., Liu X., Zhu L., Zhang H., Song X., Yang C., Sun T., Cao H, & Wang B. (2020). Cartilage Oligomeric Matrix Protein promotes epithelial-mesenchymal transition by interacting with Transgelin in Colorectal Cancer. Theranostics, 10(19), 8790-8806.

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Culturing CRC Cell Lines

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The CRC cell lines SW620, SW480, HCT-8, HCT-8/5-FU and LoVo were purchased from Keygen Biotech Co. Ltd. (Nanjing, China). SW620 and SW480 were cultured in Leibovitz’s L-15 (Gibco, Grand Island, NY) medium contained 10% inactivated fetal bovine serum (Gibco, Grand Island, NY). HCT-8, HCT-8/5-FU and LoVo cells were cultured in in RPMI 1640 (Gibco, Grand Island, NY) medium contained 10% inactivated fetal bovine serum. To develop the 5-FU resistant LoVo cells, 5-FU was added to the medium by stepwise increasing concentrations for over 6 months. HCT-8/5-FU and LoVo/5-FU were maintained in medium supplemented with 124.5 μM and 113.0 μM 5-FU. CRC cells were both incubated at 37 °C with a humidified atmosphere containing 5% CO₂. All cells lines were routinely tested for mycoplasma, which were shown to be negative.

Liu B., Liu Q., Pan S., Huang Y., Qi Y., Li S., Xiao Y, & Jia L. (2019). The HOTAIR/miR-214/ST6GAL1 crosstalk modulates colorectal cancer procession through mediating sialylated c-Met via JAK2/STAT3 cascade. Journal of Experimental & Clinical Cancer Research : CR, 38, 455.

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Evaluating Twist1 and ABCB1 in Colon Cancer

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The normal human colon mucosal epithelial cell line (NCM460), the low invasive human colon adenocarcinoma cell line (HT-29), the highly invasive human colon cancer cells (HCT-8), HCT-8/V and Bel7402 hepatocarcinoma cell line were purchased from KeyGEN BioTECH (Nanjing, China). All the cell lines were verified through short tandem repeat analysis. These cell lines were maintained in RPMI 1640 supplemented with 10% fetal bovine serum (FBS) (Hyclone, USA) in a humidified atmosphere of 5% CO₂ at 37°C (Supplementary Table 1). The HCT-8/V cell line was preserved in 1.6 mg/L vincristine.
The cells (1×10⁶) were planted in a 60 mm dish. After the cells were incubated overnight, they were transfected with pcDNA3-Twist1, pcDNA3-negative, Twist1 siRNA, and control siRNA in serum-free Opti-MEM medium (Santa Cruz, California, USA) using Lipofectamine 2000 (Invitrogen, California, USA) according to the manufacturer’s instructions. The cells were collected after 48 h for the other experiments. To test for rescue of Twist 1 silencing, pcDNA3-ABCB1 vectors were co-transfected together with the Twist1 siRNA. For EMT induction, the cells were treated with 3 ng/mL of rh-TGF-β1 for 96 hours. The neutralizing TGF-β1 antibody was used to inhibit the TGF--β1 activity.

Liu Y.R., Liang L., Zhao J.M., Zhang Y., Zhang M., Zhong W.L., Zhang Q., Wei J.J., Li M., Yuan J., Chen S., Zong S.M., Liu H.J., Meng J., Qin Y., Sun B., Yang L., Zhou H.G., Sun T, & Yang C. (2017). Twist1 confers multidrug resistance in colon cancer through upregulation of ATP-binding cassette transporters. Oncotarget, 8(32), 52901-52912.

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