Caco-2 intestinal epithelial cells (cat#: HTB-37) and HT-29 cells (cat#: HTB-38) were purchased from the American Type Culture Collection (Manassas, VA, USA). The cells were seeded at a concentration of 5 × 105 in T-75 flasks in DMEM-based CGM. The medium was changed every 2 days, and the cells were subcultured when they reached 80% confluency. To induce the differentiation of HT-29 cells into mucus-secreting goblet cells (HT-29-MTX), HT-29 cells were seeded at a concentration of 5 × 105 in T-75 flasks for 5 days, incubated with 0.1 µM MTX (Sigma-Aldrich; cat#: SIA-M9929) for 28 days, and cultured in DMEM-based CGM without MTX for 48 h [21 (link)]. The medium was replaced every 2 days. After differentiation, HT-29-MTX cell lines were harvested and stored in liquid nitrogen until use. For cell proliferation investigations, the HT-29-MTX cells were cultured in CGM at a concentration of 5 × 105 cells in T-75 flasks. After 4–5 days, 80% confluence was achieved.
Caco 2 intestinal epithelial cells
Manufactured by American Type Culture Collection
Sourced in United States
Caco-2 intestinal epithelial cells are an immortalized cell line derived from a human colorectal adenocarcinoma. These cells spontaneously differentiate in culture to form polarized monolayers that exhibit many morphological and functional characteristics of the intestinal epithelium.
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Lab products found in correlation
Penicillin, by Thermo Fisher Scientific (2 mentions)
Streptomycin, by Thermo Fisher Scientific (2 mentions)
Ht 29, by Merck Group (1 mentions)
Ht 29 cell, by American Type Culture Collection (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Fetal bovine serum (fbs), by Merck Group (1 mentions)
Penicillin streptomycin p s, by Thermo Fisher Scientific (1 mentions)
Dulbecco s modified eagle medium dmem, by Merck Group (1 mentions)
Non essential amino acids (neaa), by Thermo Fisher Scientific (1 mentions)
Kku 213, by Japanese Collection of Research Bioresources Cell Bank (1 mentions)
Ht 29, by American Type Culture Collection (1 mentions)
Minimum essential medium (mem), by Merck Group (1 mentions)
Mouse collagen 4, by Bio-Techne (1 mentions)
Giemsa stain, by Muto Pure Chemicals (1 mentions)
6 protocols using caco 2 intestinal epithelial cells
1
Differentiation of HT-29 Cells into Goblet Cells
2
Evaluating Intestinal Barrier Function
3
Caco-2 and RAW264.7 cell culture protocol
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Caco-2 intestinal epithelial cells and RAW264.7 monocytes were purchased from the American Type Culture Collection (ATCC). Both cells were maintained in Dulbecco's Modified Eagle's Medium (DMEM) supplemented with 10% heat-inactivated FBS (FBS; Gibco BRL, Grand Island, NY, USA), 100 U/mL penicillin (Thermo, Wilmington, DE, USA), and 100 U/mL streptomycin (Thermo, Wilmington, DE, USA) under 5% CO2 at 37°C. LPS (Cell Signalling, Danvers, MA, US) and probiotics combined with FVP (3:1) (Lactobacillus Enzyme Plus Powder, Hangzhou Yosto Cosmetics Co., Ltd, Hangzhou, China; TCI CO., Ltd., Taipei, Taiwan) were obtained for cell proliferation and in vitro studies.
4
Caco-2 Intestinal Epithelial Cell Culture
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Human Caco-2 intestinal epithelial cells used in the experiments were obtained from American Type Culture Collection ((ATCC), HTB-37, Germany). The passages of the Caco-2 cell line used in the microfluidics studies ranged from the 40th to 50th passages, and for the Transwell studies, passages in the range of 40th to 65th were used. The Caco-2 cells were cultured routinely in Dulbecco’s Modified Eagle Medium (DMEM; Sigma, Denmark). The culture medium is supplemented with 10% (v/v) heat-inactivated fetal bovine serum (FBS; Sigma, Denmark), 1% (v/v) nonessential amino acids (NEAA; Gibco, Denmark) and 1% (v/v) penicillin-streptomycin (P/S; Gibco, Denmark).
5
Culturing Diverse Cell Lines for Experimental Investigations
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Caco-2 intestinal epithelial cells were obtained from the American Type Culture Collection (ATCC, VA) and grown 14 days post-confluence in Eagle's Minimum Essential Medium (EMEM) supplemented with non-essential amino acids, 1 g L–1d -glucose, 2 mM l -glutamine, 10% (v/v) fetal bovine serum, and 100 U mL–1 penicillin and streptomycin. KKU-213 and KKU-213L5 cholangiocarcinoma cells were obtained from the Japanese Collection of Research Bioresources Cell Bank (JCRB, Osaka, Japan) and grown in Ham's F12 medium supplemented with non-essential amino acids, 1.8 g L–1d -glucose, 1 mM l -glutamine, 10% (v/v) fetal bovine serum, and 100 U mL–1 penicillin and streptomycin. PNT2 prostate epithelial cells were grown in RPMI-1640 medium supplemented with non-essential amino acids, 2 g L–1d -glucose, 2 mM l -glutamine, 10% (v/v) fetal bovine serum, and 100 U mL–1 penicillin and streptomycin. HT-29 colorectal epithelial cells were obtained from ATCC and grown 7 days post-confluence in McCoy's 5A medium supplemented with non-essential amino acids, 3 g L–1d -glucose, 1.5 mM l -glutamine, 10% (v/v) fetal bovine serum, and 100 U mL–1 penicillin and streptomycin. All cells were maintained at 37 °C with 5% CO2 in a humidified incubator.
6
Visualizing Probiotic Adhesion to Caco-2 Cells
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Caco-2 intestinal epithelial cells (American Type Culture Collection) were seeded for 2 weeks on minimum essential medium (MEM; Sigma, St. Louis, MO, USA) supplemented with 10% fetal bovine serum (FBS), 1% glutamine and 1% penicillin/streptomycin (100 U/mL) at 37 °C in 5% CO2 and moisture to allow cellular differentiation. Caco-2 cells were then seeded onto 20 mm coverslip coated with mouse collagen IV (Trevigen, Gaithersburg, MA, USA) in 12-well plate at 105 cells per well 24 h before being challenged with the bacteria. The adhesion of probiotics strains to Caco-2 cells was evaluated as previously described [21 (link)]. Briefly, the stationary phase lactic acid bacteria (LAB) strains grown in MRS were diluted in MEM to an optical density of 0.9 at 600 nm. An aliquot of this culture was used to challenge a confluent Caco-2 intestinal epithelial cells at a multiplicity of infection of ~100 bacteria to epithelial cell in triplicate. The binding studies were performed at 37 °C, 5% CO2 and 80% humidity for 1 h. For direct visualization of adherent bacteria, the cells were fixed in 99.8% methanol for 10 min at room temperature then stained with Giemsa stain (Muto Pure Chemicals, Tokyo, Japan) at 1:10 dilution for 20 min. A total of 100 cells were examined under the light microscope and the number of adhered bacteria to each cell was counted in 20 randomly selected fields.
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