The CCD-18 Co cell line (ATCC® CRL-1459™) was purchased from ATCC: The Global Bioresource Center (10801 University Blvd. Manassas, VA 20110, USA). The CCD-18 Co cell line was cultured using ATCC-formulated Eagle’s Minimum Essential Medium with L-glutamine (catalog No. 30-2003). To make the complete growth medium, fetal bovine serum was added to a final concentration of 10%. Every 2–3 days, a new medium was used. The cells obtained from the patient were normal myofibroblasts in the colon. The biological safety of the CCD-18 Co cell line has been classified by the American Biosafety Association (ABSA) as level 1 (BSL-1). The Caco-2 cell line was also purchased from ATCC and cultured according to the ATCC protocols. The Caco-2 cell line was obtained from a patient—a 72-year-old Caucasian male diagnosed with colon adenocarcinoma. The biological safety of the obtained material is classified as level 1 (BSL-1). To complete the medium, it was based on Eagle’s Minimum Essential Medium with L-glutamine, with the addition of fetal bovine serum to a final concentration of 20%. The medium was renewed once or twice a week.
Atcc crl 1459
Manufactured by American Type Culture Collection
Sourced in United States
ATCC CRL-1459 is a cell line derived from the human cervical adenocarcinoma HeLa cell line. It is a widely used model system for various cell biology and biomedical research applications.
Automatically generated - may contain errors
Lab products found in correlation
Caco 2 cell line, by American Type Culture Collection (2 mentions)
Atcc ccl 248, by American Type Culture Collection (2 mentions)
Ccd 18co cell line, by American Type Culture Collection (1 mentions)
Caco 2 atcc htb 37, by American Type Culture Collection (1 mentions)
Caco 2, by American Type Culture Collection (1 mentions)
Minimum essential medium (mem), by Thermo Fisher Scientific (1 mentions)
Ccd 18co, by Thermo Fisher Scientific (1 mentions)
Fetal bovine serum (fbs), by Biowest (1 mentions)
Atcc htb 38, by American Type Culture Collection (1 mentions)
Andro, by Merck Group (1 mentions)
Bx41 light microscope, by Olympus (1 mentions)
Leibovitz s l 15 medium, by Thermo Fisher Scientific (1 mentions)
Caco 2, by RIKEN BioResource Center (1 mentions)
11 protocols using atcc crl 1459
1
Culturing Colon Cell Lines CCD-18 Co and Caco-2
2
Culturing and Characterizing Colon Cell Lines
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CCD-18 Co cell line (ATCC® CRL-1459™) was purchased from ATCC: The Global Bioresource Center. CCD-18 Co cell line was cultured using ATCC-formulated Eagle’s minimum essential medium with L-glutamine (catalog No. 30-2003). To make the complete growth medium, fetal bovine serum was added to a final concentration of 10%. Every two to three days, a new medium was used. The cells obtained from the patient are normal myofibroblasts in the colon. The biological safety of the CCD-18 Co cell line has been classified by the American Biosafety Association (ABSA) as level 1 (BSL-1). The CaCo-2 cell line was also purchased from ATCC and cultured according to the ATCC protocols. The CaCo-2 cell line was obtained from a patient—a 72-year-old Caucasian male diagnosed with colon adenocarcinoma. The biological safety of the obtained material is classified as level 1 (BSL-1). To make the medium complete, Eagle’s minimum essential medium with L-glutamine had fetal bovine serum added to it to a final concentration of 20%. The medium was renewed once or twice a week.
3
Culturing Colon CCD-18Co Myofibroblast Cells
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As a cellular model, we used the colon CCD-18Co myofibroblast cell line (ATCC® CRL-1459) from the American Type Culture Collection (ATCC, Rockville, MD, USA). According to the ATCC recommendations, the culture medium selected was EMEM (pH 7.2–7.4) enriched with FBS (10% v/v) and supplemented with antibiotics (streptomycin and penicillin at 100 mg/mL and 100 U/mL, respectively), 1.5 g/L sodium bicarbonate, 1 mM sodium pyruvate, 0.1 mM nonessential amino acids, and 2 mM L-glutamine. The growth and maintenance of the cells were carried out as follows: cells seeded at 6000 cells/cm2 in T75-flasks incubated at 37 °C in incubators set with optimum growth conditions (constant humidity and 5% CO2/95% air atmosphere) for 4–5 days. At confluences ≥ 80%, the cells were subcultured and seeded at concentrations optimized for maintenance or treatments. The population doubling levels (PDL) range used in all experiments was from 26 to 32.
4
Cultivation of Colon Cell Lines CCD-18Co and Caco-2
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CCD-18Co (ATCC ® CRL-1459™) and CaCo-2 (ATCC ® HTB-37™) cell line was purchased from ATCC: The Global Bioresource Center. CCD-18Co cell line was cultured using ATCCformulated Eagle's Minimum Essential Medium with L-glutamine (catalog No. . To make the complete growth medium, fetal bovine serum was added to a final concentration of 10%. Every 2-3 days, a new medium was used. The cells obtained from the patient are normal myofibroblasts in the colon. The biological safety of the CCD-18Co cell line has been classified by the American Biosafety Association (ABSA) as level 1 (BSL-1). The CaCo-2 cell line was also cultured according to the ATCC protocols. The CaCo-2 cell line was obtained from a patient -a 72-year-old Caucasian male diagnosed with colon adenocarcinoma. The biological safety of the obtained material was classified as level 1 (BSL1). To make the medium complete we based on Eagle's Minimum Essential Medium with L-glutamine, with addition of a fetal bovine serum to a final concentration of 20%. The medium was renewed once or twice a week.
5
Culturing Cancer and Fibroblast Cells
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The T84, HCT 116 human colon cancer cell line (ATCC® CCL-248™) and the CCD-18Co human fibroblast cell line (ATCC® CRL-1459™) were purchased from the American Type Culture Collection (Manassas, VA). The T84 cells and HCT 116 cells were grown in DMEM/F-12 and RPMI-1640 nutrient media with 5% and 10% FBS respectively, and CCD-18Co cells were cultured in DMEM with 10% FBS and a 1 to 100 dilution of 100X stock MEM nonessential amino acids as previously described. [24 (link)] Bone marrow derived macrophages (BMDM) were derived from mice as previously described [17 (link)] and cultured in complete RPMI media supplemented with 10% FBS and 20% conditioned media from LADMAC cells as a source of CSF-1. [26 ] All media were supplemented with a 1X solution of antimicrobial reagents (10,000 U/ml penicillin, 10,000 μg/ml streptomycin, 25 μg/ml amphotericin B).
6
Normal Human Colon Fibroblast Cell Culture
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The normal human colon fibroblast CCD-18Co cell line (ATCC CRL-1459) was obtained from the American Type Culture Collection (ATCC; Manassas, VA, USA). CCD-18Co cells were grown in Minimum Essential Medium (MEM; Gibco, Grand Island, NY, USA) supplemented with 10% fetal bovine serum (FBS; Biowest, Nuaillé, France) and 1% 100x Antibiotic-Antimycotic solution (Nacalai Tesque, Kyoto, Japan). All cells were between passages 3–5 for all experiments and maintained at 37°C with 5% CO2.
7
Expansion and Banking of Cell Lines
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HT-29 (passages 10-20, ATCC® HTB-38™) and WiDr (passages 5-20, ATCC® CCL-218™) human colorectal adenocarcinoma cells, CCD-18Co (passages 2–15, ATCC® CRL-1459™) human colon fibroblast cells and CaCo2 (passages 12–20, ATCCR® HTB-37™) human colorectal adenocarcinoma cells differentiated in intestinal cells as described in the next section, were obtained from the American Type Culture Collection (ATCC, LGC Standards S.r.l, Italy). HaCaT cells (passages 3-7) immortalized human keratinocytes were acquired from Voden medical (Meda, MB, Italy). All the cell lines were certified by STRA and cultured as recommended.
All the cell lines were immediately expanded after delivery (up to 6 × 107 cells) and frozen down (1 × 106/vial) such that all the cell lines could be restarted after a maximum of 10 passages every 3 months from a frozen vial of the same batch of cells. Control of mycoplasma was done from a frozen vial.
All the cell lines were immediately expanded after delivery (up to 6 × 107 cells) and frozen down (1 × 106/vial) such that all the cell lines could be restarted after a maximum of 10 passages every 3 months from a frozen vial of the same batch of cells. Control of mycoplasma was done from a frozen vial.
8
Colon Cancer Cell Lines Treated with Andro
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The T84 (ATCC® CCL-248™) and COLO 205 (ATCC® CCL-222™) human colon cancer cell lines, normal colon epithelial cells FHC (ATCC®CRL-1831)TM and the CCD-18Co human fibroblast cell line (ATCC® CRL-1459™), were purchased from the American Type Culture Collection (Manassas, VA) and grown in the recommended complete tissue culture media. Cells were grown until approximately 75% confluent and then the media was replaced with fresh serum-free media containing 1/3 the concentration of antibiotic-antifungal solution (100X stock solution used at 0.33X) for 3 h. The media was then replaced with media containing 2% FBS with Andro (Sigma Aldrich, St. Louis, MO) at IC50 for 24 and 48 h. Stock Andro was prepared in DMSO and control wells received DMSO at a final concentration of 0.01%. Tunicamycin (2 μg/ml) was used as a positive control for ER stress induction. The inhibitor 4-PBA (100 mM) was used to block ER stress and NAC (20 mM) was used to block ROS. When these inhibitors were used, they were added 1 h before Andro administration.
9
Culturing Human Intestinal Fibroblasts
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Human intestinal CCD-18Co fibroblast cell line (ATCC CRL-1459) was obtained from American Type Culture Collection (ATCC, Georgetown, DC, USA) and then cultured in DMEM that was supplemented with 10% of Fetal Bovine Serum (FBS), 100 U/ml penicillin, 100 mg/ml streptomycin, and 2 mM glutamine. The cell cultures were maintained in a humidified atmosphere of 95% air and 5% CO2 at 37 °C. Where not otherwise specified, the reagents for cell biology and consumables were purchased from EuroClone (EuroClone, West York, UK). After reaching 80% confluence, the cells were sub-cultured; the experiments were carried out at the 15th passage.
10
Evaluating B. pullicaecorum Effects on CRC Cells
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SW480 colon cancer cell line (ATCC CRL-1459; AJCC stage II) and SW620 lymph node metastatic derivative cell line (ATCC CRL-1831; AJCC stage III) from the same patient were purchased from American Type Culture Collection. The two cell lines were expanded in complete medium [Leibovitz's L-15 medium (Thermo Fisher Scientific, Inc.) supplemented with 10% fetal bovine serum (NQBB International Biological Co., Ltd.)] under 100% atmospheric air (without CO2) in a 37°C humidified incubator. To evaluate the effects of B. pullicaecorum on CRC cell proliferation, the numbers of SW480 and SW620 cells were respectively counted at days 1 and 5. Briefly, cells incubated with or without 10% conditioned medium from B. pullicaecorum cultures in complete medium were imaged under an Olympus BX41 light microscope (Olympus Corporation) and analyzed using ImageJ software (version 1.52; National Institutes of Health) (34 (link)).
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