Atcc crl 1739

Manufactured by American Type Culture Collection

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ATCC CRL 1739 is a cell line derived from human fetal lung fibroblasts. It is a commonly used in vitro model for cellular and molecular studies.

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15 protocols using atcc crl 1739

Generation and Characterization of hiPSC-CMs

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The human-induced pluripotent stem cells (hiPSCs) were generated from primary fibroblasts derived from skin biopsies of three healthy donors. The generation of hiPSC-CMs from hiPSCs have been described in our previous studies (14 (link), 15 (link)). The successfully differentiation of hiPSC-CMs was confirmed by immunofluorescence of cardiac markers α-actinin and myosin light chain 4 (Myl4) (Supplementary Figure 1) together with spontaneous cell beating and cardiac action potential features.
The two cancer cell lines AGS (ATCC® CRL-1739™) and SW480[SW-480] (ATCC® CCL-228™) were bought from ATCC (LGC Standards GmbH, Wesel, Germany). Cancer cells (1 × 10⁵ cells) were seeded in T75 flasks, and then cultured in DMEM (10 mL) supplemented with 10% FBS and 1% Pen/Strep. According to the growth curve of the cell lines by cell counting and MTT assay (Supplementary Figure 2), we harvested the cancer cell media on day 8 at the end of the logarithmic growth phase with stable cancer cell number and secretion. PCR data showed that different concentrations of the fresh cancer cell media had no relevant effects on the ion channel gene expressions of hiPSC-CMs (Supplementary Figure 3). Since the medium cultured with cancer cells showed concentration-dependent effects (Supplementary Figure 4), we chose the concentration of 20% of cancer cell media for the further experiments.

Zhong R., Zhang F., Yang Z., Li Y., Xu Q., Lan H., Lang S., Cyganek L., Burgermeister E., El-Battrawy I., Zhou X., Akin I, & Borggrefe M. (2022). Regulation of Ion Channel Function in Human-Induced Pluripotent Stem Cell-Derived Cardiomyocytes by Cancer Cell Secretion Through DNA Methylation. Frontiers in Cardiovascular Medicine, 9, 839104.

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Isolation and Culture of Cancer Cell Lines

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The LoVo cell line was isolated from a metastatic nodule by Dolfini et al (53 (link)); its MDR variant, LoVo DX, and a gastric adenocarcinoma cell line (AGS) were used in this study. The MDR cell line, LoVo DX, was obtained by the prolonged culture of drug-sensitive parental LoVo WT cells in medium containing DOX (Adriblastina; Pharmacia & Upjohn, Milan, Italy) as previously described by Grandi et al (54 (link)). LoVo DX cells are also resistant to other drugs, such as etoposide and vincristine (39 (link),53 (link)). The LoVo cells were a kind gift from Professor E. Dolfini (University of Milan, Milan, Italy). The AGS cell line (homo sapiens gastric adenocarcinoma) was obtained from the American Type Culture Collection (ATCC^® CRL-1739™; ATCC, Manassas, VA, USA). All cell lines were grown in Ham’s F-12 medium containing L-glutamine supplemented with 10% FBS, 1% minimum essential medium (MEM) vitamins, 1% MEM non-essential amino acid, penicillin (100 U/ml) and streptomycin (100 µg/ml) and were incubated in a humidified atmosphere of 5% CO₂ in a water-jacketed incubator at 37°C. For each passage, exponentially growing LoVo and AGS cells were harvested with 10 mM EDTA in PBS and then by further addition of 0.25% trypsin solution in PBS. The trypsin activity was quenched by the addition of complete F-12 medium.

Ohkubo S., Mancinelli R., Miglietta S., Cona A., Angelini R., Canettieri G., Spandidos D.A., Gaudio E, & Agostinelli E. (2019). Maize polyamine oxidase in the presence of spermine/spermidine induces the apoptosis of LoVo human colon adenocarcinoma cells. International Journal of Oncology, 54(6), 2080-2094.

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Hummingbird Phenotype Induction by H. pylori

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For H. pylori-induced hummingbird phenotype analysis, AGS cells were cultured in RPMI 1640 medium containing 10% FBS. Each of the H. pylori isolates were suspended in RPMI 1640 and this suspension was used for AGS cell infection at a multiplicity of infection (MOI) of 100 for 5 h. The morphology of the AGS cells was determined using a microscope, and cells showing an elongated hummingbird phenotype (a ratio of the longest protrusion to the shortest diameter of greater than 2) were scored in ten different fields (10 samples at x40 magnification). AGS cells (ATCC CRL 1739) were purchased from Food Industry Research and Development Institute (FIRDI), Taiwan, in 2002.

Chang C.C., Kuo W.S., Chen Y.C., Perng C.L., Lin H.J, & Ou Y.H. (2016). Fragmentation of CagA Reduces Hummingbird Phenotype Induction by Helicobactor pylori. PLoS ONE, 11(3), e0150061.

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Helicobacter pylori Infection in AGS Cells

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H. pylori reference strain 26695 (American Type Culture Collection [ATCC] 700392) and AGS human gastric adenocarcinoma cell line (ATCC CRL-1739) were purchased from the ATCC (Manassas, VA, USA). H. pylori was grown on tryptic soy agar (TSA) supplemented with 5% sheep blood (Dr. Plate Biotech, Taipei, Taiwan) under microaerophilic conditions obtained using an anaerobic BD GasPak (BD Biosciences, Franklin Lakes, NJ, USA) at 37°C for 48–72 h. LR-JB3 isolated from a dairy product (6 (link)) was cultured in De Man, Rogosa, and Sharpe (MRS) broth (BD Biosciences, Franklin Lakes, NJ, USA) at 37°C for 12–18 h. AGS cells were cultured in Roswell Park Memorial Institute 1640 (RPMI) medium (Gibco-Life Technologies, Gaithersburg, MD, USA) supplemented with 10% fetal bovine serum (FBS) (Sigma-Aldrich, St. Louis, MO, USA), 100 μg/mL of penicillin, and 100 μg/mL of streptomycin (Thermo Scientific, Waltham, MA, USA) at 37°C and 5% of CO₂. During infection experiments, AGS cells were washed three times using phosphate buffer solution (PBS), and the culture media were replaced with antibiotic-free RPMI media containing 10% FBS for further incubation.

Do A.D., Su C.H, & Hsu Y.M. (2022). Antagonistic Activities of Lactobacillus rhamnosus JB3 Against Helicobacter pylori Infection Through Lipid Raft Formation. Frontiers in Immunology, 12, 796177.

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Gastric Adenocarcinoma Cell Culture

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Gastric adenocarcinoma cells (AGS) (ATCC CRL-1739) were obtained from ATCC (Manassas, VU, USA). Cells were grown in RPMI medium (GIBCO, Grand Island, NY, USA) with the following supplements: 10% heat-inactivated fetal bovine serum (Euroclone, Pero, Italy), 2 mM of L-glutamine, 50 ng/mL of streptomycin, and 50 units/mL of penicillin, all purchased from Sigma-Aldrich (St. Louis, MO, USA). Cells were maintained at 37 °C in a humidified 5% CO₂ atmosphere.

Esposito T., Pisanti S., Mauro L., Mencherini T., Martinelli R, & Aquino R.P. (2023). Activity of Colocasia esculenta (Taro) Corms against Gastric Adenocarcinoma Cells: Chemical Study and Molecular Characterization. International Journal of Molecular Sciences, 25(1), 252.

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Culturing AGS Gastric Cancer Cells

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The human gastric epithelial cell line gastric adenocarcinoma (AGS, American Type Culture Collection (ATCC) CRL-1739) was purchased from ATCC (Rockville, MD, USA). The cells were grown in a complete medium consisting of RPMI (Roswell Park Memorial Institute) 1640 (Gibco, Grand Island, NY, USA) supplemented with 10% heat-inactivated fetal bovine serum (Gibco), 2 mM glutamine, 100 U/mL penicillin, and 100 µg/mL streptomycin (Sigma-Aldrich, St. Louis, MO, USA). The cells were cultured at 37 °C in a humidified atmosphere of 95% air and 5% CO₂.

Lee H., Lim J.W, & Kim H. (2020). Effect of Astaxanthin on Activation of Autophagy and Inhibition of Apoptosis in Helicobacter pylori-Infected Gastric Epithelial Cell Line AGS. Nutrients, 12(6), 1750.

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H. pylori Infection in AGS Cells

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The human gastric epithelial AGS cells (gastric adenocarcinoma, ATCC CRL 1739; American Type Culture Collection, Rockville, MD, USA) were grown as described previously [18 (link)]. H. pylori cells (NCTC 11637, American Type Culture Collection) were grown on chocolate agar plates (Becton Dickinson Microbiology Systems, Cockeysville, MD, USA) at 37°C, under microaerophilic conditions, using an anaerobic chamber (BBL Campy Pouch^® System; Becton Dickinson Microbiology Systems, Franklin Lakes, NJ, USA). The H. pylori strain NCTC 11637 expresses the proteins of CagA, VacA, cytotoxin and urease [19 (link)].
AGS cells were seeded overnight to reach 80% confluency. The H. pylori was harvested and suspended in antibiotic-free RPMI 1640 medium supplemented with 10% FBS, and then added to the AGS cell culture at a 50 : 1 ratio of bacterium : AGS cells. AGS cells (1.0 × 10⁵/mL) were pre-treated with 5 or 10 μM β-carotene (dissolved in tetrahydrofuran [17 ]) for 2 hours prior to the addition of the H. pylori. The infected cells were incubated for an additional 6 hours for determination of the levels of p-GSK3β, GSK3β, and β-catenin or 24 hours for determination of cell viability and the levels of c-myc and cyclin E) in the presence of H. pylori. The 5 and 10 μM β-carotene concentrations used in the pre-treatment are reportedly nontoxic [20 (link),21 (link)].

Kim D., Lim J.W, & Kim H. (2019). β-carotene Inhibits Expression of c-Myc and Cyclin E in Helicobacter pylori-infected Gastric Epithelial Cells. Journal of Cancer Prevention, 24(3), 192-196.

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Astaxanthin Inhibits H. pylori Infection

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The human gastric epithelial AGS cells (gastric adenocarcinoma, ATCC CRL 1739; American Type Culture Collection, Rockville, MD, USA) were grown in RPMI 1640 medium (GIBCO, Grand Island, NY, USA) supplemented with 10% FBS, 2 mM glutamine, 100 U/mL penicillin, and 100 μg/mL streptomycin (Sigma-Aldrich) and cultured at 37°C under 95% air and 5% CO₂.
H. pylori bacteria (NCTC 11637; American Type Culture Collection) were grown on chocolate agar plates (Becton Dickinson Microbiology Systems, Cockeysvile, MD, USA) at 37°C, under microaerophilic conditions, using an anaerobic chamber (BBL Campy Pouch^® System; Becton Dickinson Microbiology Systems, Franklin Lakes, NJ, USA).
AGS cells were seeded overnight to reach 80% confluency. The H. pylori was harvested and suspended in antibiotic-free RPMI 1640 medium supplemented with 10% FBS, and then added to the AGS cell culture at the ratio of bacterium: AGS cells, 50: 1. AGS cells (1.0 × 10⁵/mL) were pre-treated with astaxanthin (at final concentration of 1 and 5 μM) for 3 hours before adding the H. pylori and cultured for 1 hour.

Kim S.H., Lim J.W, & Kim H. (2019). Astaxanthin Prevents Decreases in Superoxide Dismutase 2 Level and Superoxide Dismutase Activity in Helicobacter pylori-infected Gastric Epithelial Cells. Journal of Cancer Prevention, 24(1), 54-58.

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Culturing Human Gastric Adenocarcinoma AGS Cells

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Human gastric adenocarcinoma cell-line AGS cells ATCC® CRL-1739™ (American Type Culture Collection, Rockville, MD, USA), were routinely cultured in Ham's F-12 K medium (Gibco, Waltham, Massachusetts,USA) supplemented with 1% (v/v) Penstrep, 1% (v/v) glutamine and 10% (v/v) fetal calf serum (Gibco) at 37 °C and 5% CO₂.

D'ambrosio S., Ventrone M., Fusco A., Casillo A., Dabous A., Cammarota M., Corsaro M.M., Donnarumma G., Schiraldi C, & Cimini D. (2022). Limosilactobacillus fermentum from buffalo milk is suitable for potential biotechnological process development and inhibits Helicobacter pylori in a gastric epithelial cell model. Biotechnology Reports, 34, e00732.

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Cultivation of Gastric Adenocarcinoma Cells

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Gastric epithelial AGS cells (gastric adenocarcinoma, ATCC CRL 1739, Rockvile, MD, USA) were purchased from the American Type Culture Collection (Rockvile, MD, USA). The cells were grown in complete medium, consisting of RPMI 1640 medium (GIBCO, grand Island, NY, USA), supplemented with 10% fetal bovine serum (GIBCO), 2 mM glutamine, 100 U/mL penicillin, and 100 μg/mL streptomycin (Sigma-Aldrich, St. Louis, MO, USA). The cells were cultured at 37 °C under a humidified atmosphere of 95% air and 5% CO₂.

Park Y., Lee H., Lim J.W, & Kim H. (2019). Inhibitory Effect of β-Carotene on Helicobacter pylori-Induced TRAF Expression and Hyper-Proliferation in Gastric Epithelial Cells. Antioxidants, 8(12), 637.

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