Ags cells

Manufactured by American Type Culture Collection

Sourced in United States

AGS cells are a type of human gastric adenocarcinoma cell line. They are commonly used in research as an in vitro model for the study of gastric cancer biology.

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51 protocols using ags cells

AGS Cells Infection by H. pylori

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AGS cells (ATCC, Manassas, VA) were cultured in DME/F12 medium containing 10% heat inactivated FBS, 100 units/ml penicillin, and 100 µg/ml streptomycin (Invitrogen, Carlsbad, CA) as described [14 (link)]. H. pylori strain 26695 was maintained on Columbia blood agar containing 7% defibrinated horse blood (Cleveland Scientific, Bath, OH), 20 µg/ml bacitracin, 20 µg/ml trimethoprim, 16 µg/ml cefsulodin, 6.0 µg/ml vancomycin, and 2.5 µg/ml fungizone (Sigma, St. Louis, MO) under microaerophilic conditions as described [14 (link)]. AGS cells were untreated or treated for 1 hr or 4 hr with H. pylori at a multiplicity of infection (MOI)=100. In selected experiments, AGS cells were treated for 72 hr with 1.0 µM of 5-aza-dC (Sigma) in the absence of H. pylori.

Guang W., Czinn S.J., Blanchard T.G., Kim K.C, & Lillehoj E.P. (2014). Genetic Regulation of MUC1 Expression by Helicobacter pylori in Gastric Cancer Cells. Biochemical and biophysical research communications, 445(1), 145-150.

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AGS cells Pretreatment with WPE and H. pylori

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AGS cells were purchased from ATCC (Manassas, VA), where the cells were properly stored and routinely authenticated (including DNA fingerprinting). After resuscitation in our lab, all the cells were used no longer than 6 months. AGS cells were cultured in RPMI-1640 medium (Gibco BRL). All mediums supplemented with 10% fetal bovine serum (Gibco BRL) at 37°C in 5% CO₂. AGS cells were pretreated with WPE for 1 h and stimulated with 100 MOI H. pylori for 48 h. The control group was loaded with the same concentration of the dissolving media of DMSO.

Park J.M., Han Y.M., Lee H.J., Hwang S.J., Kim S.J, & Hahm K.B. (2021). Transcriptome profiling analysis of the response to walnut polyphenol extract in Helicobacter pylori-infected cells. Journal of Clinical Biochemistry and Nutrition, 68(3), 201-214.

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Coculture of AGS Cells with H. pylori

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AGS cells (ATCC^®) were grown in Gibco^R RPMI 1640 medium (Life Technologies, Pasching, Austria) supplemented with 10% fetal bovine serum (FBS). Approximately 16 h prior to the experiment, the cell culture medium was replaced with fresh RPMI 1640 supplemented with 0.2% FBS. The CagA-expressing H. pylori strain P1 wild-type [11 (link), 68 (link)] was cultured on GC agar supplemented with horse serum, vancomycin, trimethoprim, nystatin and vitamins in microaerophilic conditions for 48–72 h, and then a bacterial suspension in PBS was prepared. H. pylori was added to AGS cells at a multiplicity of infection of 100. Recombinant human TNFα and IL-1β (R&D Systems, Minneapolis, MN, USA) were used at a final concentration of 10 ng/ml.

Sokolova O., Kähne T., Bryan K, & Naumann M. (2018). Interactome analysis of transforming growth factor-β-activated kinase 1 in Helicobacter pylori-infected cells revealed novel regulators tripartite motif 28 and CDC37. Oncotarget, 9(18), 14366-14381.

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Kimchi Pretreatment Inhibits H. pylori in AGS Cells

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AGS cells were purchased from ATCC (Manassas, VA), where the cells were properly stored and routinely authenticated (including DNA fingerprinting). After resuscitation in our lab, all the cells were used no longer than 6 months. AGS cells were cultured in RPMI-1640 medium (Gibco BRL, Gaithersburg, MD). All mediums supplemented with 10% fetal bovine serum (Gibco BRL) at 37°C in 5% CO₂. AGS cells were pretreated with kimchi (5 μg/ml) for 1 h and stimulated with H. pylori [100 multiplicity of infection (MOI)] for 24 h. The control group was loaded with the same concentration of the dissolving media of DMSO.

Park J.M., Han Y.M., Oh J.Y., Lee D.Y., Choi S.H, & Hahm K.B. (2021). Transcriptome profiling implicated in beneficiary actions of kimchi extracts against Helicobacter pylori infection. Journal of Clinical Biochemistry and Nutrition, 69(2), 171-187.

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Macrophage-Modulated 5FU Cytotoxicity

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AGS cells (ATCC, Rockville, USA) and THP-1 cells (ATCC, Rockville, USA) were plated in 96-well plates at a final concentration of 1 × 10⁴ cells/well for 24 h before the experiments. Cells were then treated with various concentrations of 5FU for 24 h with (or without) conditioned media (CM) from M1 and M2 macrophages for 24 h. After treatment, the media in the wells were removed and replaced by fresh prewarmed media. WST-1 was added to each well and the plates were incubated at 37°C for an additional period of 4 h. The absorbance was measured using a Multiplate reader at a wavelength of 490 nm.

Ngabire D., Niyonizigiye I., Patil M.P., Seong Y.A., Seo Y.B, & Kim G.D. (2020). M2 Macrophages Mediate the Resistance of Gastric Adenocarcinoma Cells to 5-Fluorouracil through the Expression of Integrin β3, Focal Adhesion Kinase, and Cofilin. Journal of Immunology Research, 2020, 1731457.

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Synthesis and Characterization of Carbohydrate Analogs

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Organic chemicals and anti-FLAG antibodies were purchased from Millipore Sigma. H. pylori strain G27^{84 (link)} was a gift of Manuel Amieva (Stanford University). Bacterial cells (P. shigelloides ATCC 51903; V. vulnificus ATCC 43382; S. aureus ATCC BAA-1708; C. jejuni ATCC 33560; B. fragilis ATCC 23745) and AGS cells (ATCC CRL-1739) were purchased from ATCC and grown according to the supplier’s instructions. Ac₄GlcNAc (1), Ac₄GlcNAz (2), Ac₄GalNAz (3), Ac₂Bac-diNAz (4), and Phos-FLAG were synthesized as previously described.^{44 (link), 85 , 86}l-FucNAz (5), l-PneNAz (6), l-RhaNAz (7), and l-QuiNAz (8) were synthesized using standard organic chemistry procedures and characterized by standard techniques including ¹H and ¹³C NMR spectroscopy and mass spectrometry. Analogs 5–8 were purified using flash silica gel chromatography.

Luong P., Ghosh A., Moulton K.D., Kulkarni S.S, & Dube D.H. (2022). Synthesis and Application of Rare Deoxy Amino l-Sugar Analogs to Probe Glycans in Pathogenic Bacteria. ACS infectious diseases, 8(4), 889-900.

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Cell Culture Conditions for Gastric Cancer

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All the tissue culture reagents were from Invitrogen (France). AGS cells (ATCC CRL 1739) and NCI-N87 (ATCC CRL 5822) cells were routinely grown in DMEM/F12 medium, whereas MKN74 (JCRB0255), MKN28 (JCRB0253), MKN7 (JCRB1025) cells were grown in RPMI 1640 medium. Both media were supplemented with 200 mM L-glutamine and 10% heat-inactivated fetal calf serum (FCS). Cells were cultured at 37 °C in a 5% CO₂ atmosphere. Before being added directly in the culture media at the final concentration indicated in the figures, the conjugates were diluted in serum-free medium from a 50 mM stock solution in sterile water. The culture medium, with or without the conjugates, was changed every other day.

Staedel C., Tran T.P., Giraud J., Darfeuille F., Di Giorgio A., Tourasse N.J., Salin F., Uriac P, & Duca M. (2018). Modulation of oncogenic miRNA biogenesis using functionalized polyamines. Scientific Reports, 8, 1667.

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Cell Culture Protocols for Cancer Cell Lines

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AGS cells (ATCC CRL 1739) were cultured in F12 medium (Invitrogen), MKN-45 cells (JCRB0254; RIKEN Cell Bank, Japan) and HT29 cells (ATCC HTB-38; human colorectal adenocarcinoma) were cultured in DMEM (Invitrogen), COLO205 cells (CCL-222; human colon adenocarcinoma cells) were cultured in RPMI 1640 medium (Invitrogen). Cell were cultured in medium supplemented with 10% fetal bovine serum (HyClone, Logan, UT) and incubated at 37°C in a humid atmosphere containing 5% CO₂.

Lin H.J., Jiang Z.P., Lo H.R., Feng C.L., Chen C.J., Yang C.Y., Huang M.Z., Wu H.Y., Chen Y.A., Chen Y., Chiu C.H, & Lai C.H. (2019). Coalescence of RAGE in Lipid Rafts in Response to Cytolethal Distending Toxin-Induced Inflammation. Frontiers in Immunology, 10, 109.

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Bacterial Glycan Biosynthesis Protocols

Cited 1 time

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Organic chemicals
and anti-FLAG antibodies were purchased from Sigma-Aldrich. H. pylori strain G27^{58 (link)} was a gift of Manuel Amieva (Stanford University). B. fragilis (ATCC 23745) and AGS cells (ATCC CRL-1739)
were purchased from ATCC and grown according to the supplier’s
instructions. Ac₄GlcNAc, Ac₄GlcNAz, Ac₄GalNAz, and Phos-FLAG were synthesized as previously described.^{17 (link),59 (link),60 (link)} BacSBn 4, DATSBn 5, and FucSBn
6 were synthesized by using standard organic chemistry procedures
and characterized by standard techniques including ¹H and ¹³C NMR spectroscopy and mass spectrometry. Analogues 4–6
were purified using flash silica gel chromatography.

Quintana I.D., Paul A., Chowdhury A., Moulton K.D., Kulkarni S.S, & Dube D.H. (2023). Thioglycosides Act as Metabolic Inhibitors of Bacterial Glycan Biosynthesis. ACS Infectious Diseases, 9(10), 2025-2035.

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Generation of NF-κB Reporter AGS Cells

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AGS cells (CRL-1739) and HEK 293 cells (CRL-1573) were purchased from ATCC and tested for mycoplasma contamination. AGS cells stably expressing a luciferase-based NF-κB reporter were generated by transfection of the plasmid pGL4.32[luc2P/NF-κB-RE/Hygro] (Promega), antibiotic selection and cloning. AGS cells stably transfected with the NF-κB reporter were also transfected with a mix of shRNAs targeting NOD1. Colonies were selected using puromycin (10 µg/mL) and tested for NOD1 expression by real-time RT-PCR and Western blot.

Suarez G., Romero-Gallo J., Piazuelo M.B., Wang G., Maier R.J., Forsberg L.S., Azadi P., Gomez M.A., Correa P, & Peek RM J.r. (2015). Modification of Helicobacter pylori peptidoglycan enhances NOD1 activation and promotes cancer of the stomach. Cancer research, 75(8), 1749-1759.

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