The cervical cancer-derived HeLa and the HNSCC-derived UD-SCC-2 (kindly provided by Prof. Dr. H. Bier, University of Düsseldorf, Germany) cell lines were grown in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% FBS (fetal bovine serum), 2 mmol/L l -glutamine (Capricorn Scientific GmbH, Germany), 50 μg/mL gentamicin (Biochrom GmbH, Germany), 100 U/mL penicillin/streptomycin (Capricorn Scientific GmbH, Germany), and 50 μg/mL amphotericin B (Biochrom GmbH, Germany). Fresh VX2 cells were derived from a VX2 carcinoma of a New Zeeland White (NZW) rabbit. The generation of VX2 tumors in NZW rabbits was approved by the regional board Giessen, Germany (V54-19c20-15 h01 MR 20/26 Nr. 83/2015) according to the German Animal Protection Law. Transiently growing VX2 cells (max. 150 passages) were then cultured in DMEM/Ham’s F-12 media containing 2 mmol/L l -glutamine (Capricorn Scientific GmbH, Germany) supplemented with 10% FBS, 50 μg/mL gentamicin (Biochrom GmbH, Germany), 100 U/mL penicillin/streptomycin (Capricorn Scientific, Germany), and 50 μg/mL amphotericin B (Biochrom GmbH, Germany). All three cell lines were cultured at 37 °C, 5% CO2 in a humidified incubator. Cells were grown as a monolayer until reaching approximately 80% confluency.
L glutamine
Manufactured by Capricorn
Sourced in Germany, United States
L-glutamine is an amino acid that plays a crucial role in cellular metabolism. It serves as a precursor for the synthesis of other amino acids and is involved in various metabolic processes within the body.
Automatically generated - may contain errors
Lab products found in correlation
Penicillin streptomycin, by Capricorn (15 mentions)
Penicillin, by Capricorn (12 mentions)
Streptomycin, by Capricorn (12 mentions)
Fetal bovine serum (fbs), by Capricorn (8 mentions)
Phosphate buffered saline (pbs), by Capricorn (7 mentions)
Trypsin edta, by Capricorn (5 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (4 mentions)
Rpmi 1640, by Capricorn (4 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (4 mentions)
Sodium pyruvate, by Capricorn (3 mentions)
Dulbecco s modified eagle medium dmem, by Capricorn (3 mentions)
Dulbecco modified eagle medium (dmem), by Capricorn (3 mentions)
Ht 29, by American Type Culture Collection (3 mentions)
Fetal calf serum (fcs), by Merck Group (3 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (3 mentions)
Penicillin streptomycin, by Sartorius (2 mentions)
Plx4032, by MedChemExpress (2 mentions)
Ht 29, by MedChemExpress (2 mentions)
Foetal bovine serum (fbs), by PAN Biotech (2 mentions)
Fetal bovine serum (fbs), by Merck Group (2 mentions)
55 protocols using l glutamine
1
Cell Culture Protocol for Cancer Cell Lines
2
Establishment of BRAFV600E-Mutant Cell Lines
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Human colon carcinoma cell line RKO and human melanoma cell line A375 both harbouring the BRAFV600E mutation were purchased from the American Type Culture Collection (Manassas, VA, USA). Both RKO and vemurafenib-resistant RKO cells were maintained in Eagle’s Minimum Essential Medium (Capricorn Scientific, Ebsdorfergrund, Germany) supplemented with 10% fetal bovine serum (Capricorn Scientific, Germany), 2 mM L-glutamine (Capricorn Scientific, Germany), penicillin (100 U/mL) (Capricorn Scientific, Germany) and streptomycin (100 µg/mL) (Capricorn Scientific, Germany) in humified atmosphere with 5% CO2 at 37 °C. Melanoma cell line A375 and its vemurafenib-resistant counterpart were maintained in Dulbecco’s Modified Eagle Medium (Capricorn Scientific, Germany) supplemented with 10% fetal bovine serum (Capricorn Scientific, Germany), 2 mM L-glutamine (Capricorn Scientific, Germany), penicillin (100 U/mL) (Capricorn Scientific, Germany), streptomycin (100 µg/mL) (Capricorn Scientific, Germany) and 1 mM sodium pyruvate (Capricorn Scientific, Germany) in humified atmosphere with 5% CO2 at 37 °C.
3
Cytoskeletal Dynamics: Avermectin, Paclitaxel, and Colchicine
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Avermectin B1a was purchased from BioAustralis (BIA-A1010, BioAustralis, New South Wales, Australia), paclitaxel was obtained from Cytoskeleton (TXD01, Cytoskeleton, Denver, CO, USA), and colchicine was provided by Sigma Aldrich (C9754, Sigma Aldrich, St. Louis, MO, USA). Dulbecco’s Modified Eagle Medium (DMEM), the buffer Dulbecco’s PBS, and L-Glutamine (200 mM) were purchased from Capricorn (Capricorn, Dusseldorf, Germany), while fetal bovine serum (FBS), penicillin/streptomycin, and trypsin/EDTA were obtained from Biological Industries (Biological Industries, Jezreel Valley, Israel). The cell-counting dye trypan blue (0.4%) was purchased from Thermo Fisher Scientific (15250-061, Thermo Fisher Scientific, Waltham, MA, USA). Dimethyl sulfoxide (DMSO) was obtained from Carlo Erba (445106, Carlo Erba, Val de Reuil, Normandie, France).
3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) was used for cell viability assay and was purchased from Thermo Fisher Scientific (M6494, Thermo Fisher Scientific, USA). A tubulin polymerization assay was performed by using a tubulin polymerization assay kit from Cytoskeleton (BK004P, Cytoskeleton, Denver, CO, USA), whereas the apoptosis assay was performed by using annexin V-FITC/PI apoptosis kit purchased from Elabscience (E-CK-A211, Elabscience, Huston, TX, USA).
3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) was used for cell viability assay and was purchased from Thermo Fisher Scientific (M6494, Thermo Fisher Scientific, USA). A tubulin polymerization assay was performed by using a tubulin polymerization assay kit from Cytoskeleton (BK004P, Cytoskeleton, Denver, CO, USA), whereas the apoptosis assay was performed by using annexin V-FITC/PI apoptosis kit purchased from Elabscience (E-CK-A211, Elabscience, Huston, TX, USA).
4
Generating Vemurafenib-Resistant Colon Cancer Cell Lines
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Human colon carcinoma cell lines Caco-2 (BRAFwt/KRASwt), SW480 (BRAFwt/KRASG12V), HCT 116 (BRAFwt/KRASG13D) and HT-29 and RKO (BRAFV600E/KRASwt) were purchased from the ATCC and maintained in Dulbecco′s Modified Eagle′s Medium (DMEM) or Minimum Essential Medium (MEM) supplemented with 10% fetal bovine serum (FBS), 2 mM L-glutamine, penicillin (100 U/mL) and streptomycin (100 µg/mL) (Capricorn Scientific, Ebsdorfergrund, Germany) in a humified atmosphere with 5% CO2 at 37 °C.
In order to eliminate molecular features of resistance that might be cell line-specific, we developed two vemurafenib (PLX4032)-resistant colon cancer cell lines derived from HT-29 and RKO cell lines by exposing the cells to successively increasing concentrations of PLX4032 (MedChemExpress, Monmouth Junction, NJ, USA) in a period of about 6 months until a clinically relevant dose (11.52 µM) [41 (link)] was reached. Established resistance phenotypes were confirmed by the MTT assay showing an increase in the IC50 values by 8- and 10-fold in the resistant HT-29 and RKO cells, respectively, in comparison with their sensitive counterparts (Supplementary Table S5 ).
In order to eliminate molecular features of resistance that might be cell line-specific, we developed two vemurafenib (PLX4032)-resistant colon cancer cell lines derived from HT-29 and RKO cell lines by exposing the cells to successively increasing concentrations of PLX4032 (MedChemExpress, Monmouth Junction, NJ, USA) in a period of about 6 months until a clinically relevant dose (11.52 µM) [41 (link)] was reached. Established resistance phenotypes were confirmed by the MTT assay showing an increase in the IC50 values by 8- and 10-fold in the resistant HT-29 and RKO cells, respectively, in comparison with their sensitive counterparts (
5
Culturing human periodontal ligament fibroblasts and THP1 monocytes
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Human periodontal ligament fibroblasts (hPdLFs, Lonza, Basel, Switzerland) were grown in DMEM with 4.5 g/L glucose (Capricorn Scientific GmbH, Ebsdorfergrund, Germany) with 10% fetal bovine serum (FBS; Thermo Fisher Scientific, Carlsbad, CA, USA), 1% penicillin/streptomycin (Gibco, Thermo Fisher Scientific, Carlsbad, CA, USA) and 1% L-ascorbid acid (Sigma Aldrich, St. Louis, MO, USA). Cells were regularly subcultured when a confluency of 75% was reached. Subcultures four to eight were used.
THP1 monocytic cells (DSMZ, Braunschweig, Germany) were cultured in RPMI 1640 with L-glutamine and sodium bicarbonate (Capricorn Scientific GmbH, Ebsdorfergrund, Germany) supplemented with 10% FBS and 1% penicillin/streptomycin.
THP1 monocytic cells (DSMZ, Braunschweig, Germany) were cultured in RPMI 1640 with L-glutamine and sodium bicarbonate (Capricorn Scientific GmbH, Ebsdorfergrund, Germany) supplemented with 10% FBS and 1% penicillin/streptomycin.
6
Hyphal Fragment Injection Protocol for G. mellonella
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To generate hyphal fragments that could be injected into G. mellonella larvae, mycelia from F. senegalensis were obtained from 2-week old cultures grown on Sabouraud agar plates and inoculated in RPMI 1640 culture medium supplemented with L-glutamine (0.3 g/l; Capricorn-Scientific, Germany), 20 m m mopholinepropanesulfonic acid (MOPS; Sigma, USA), and chloramphenicol (100 mg/l; Oxoid, Basingstroke, UK). After 2 weeks of incubation at 37°C, hyphae were collected by filtering them through a 0.22 µm filter (Nalgene, Abcoude, The Netherlands) and washed with phosphate-buffered saline (PBS; Gibco, USA). To obtain hyphal fragments, the washed hyphae were sonicated for 10 s at 20 µm (Soniprep, Beun de Ronde, The Netherlands), as previously done for M. mycetomatis.8 (link) To establish the lethal dose, different inocula were prepared in PBS. These were: 10 mg/larvae, 4 mg/larvae, 0.4 mg/larvae, and 0.04 mg/larvae. A total of 40 µl was injected into the last left pro-leg of the larvae with an insulin 29 G U-100 needle (BD Diagnostics, Sparsk, USA). Uninfected controls were injected with 40 µl PBS. A total of 15 larvae per group were used, and every test was replicated three times. Larvae were placed in a petridish containing Whattman paper and incubated at 37°C in a normal incubator. Survival was monitored for 10 days.
7
SARS-CoV-2 Cell Culture and Titration
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Minimum Essential Medium (MEM; GIBCO, Thermo Fisher Scientific, MA USA) supplemented with penicillin (100 IU mL-1), streptomycin (100 µg mL-1), and L-glutamine (2 mM) (all from Capricorn Scientific, Germany) was used. Foetal bovine serum (PAN-Biotech, Germany) was inactivated at 56°C for 60 minutes prior the use. For the Vero E6 cell propagation and maintenance medium with 10% FBS was used. In SARS-CoV-2 titration (CCID50 assay) and SARS-CoV-2 neutralisation (ED50) assays medium was supplemented with 2.5% FBS.
8
Maintenance of Testicular Cancer Cell Lines
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The cell lines employed in this work were maintained in DMEM/Ham’s F-12 medium with L-Glutamine (Capricorn Scientific, Ebsdorfergrund, Germany), 10% FCS and 1% penicillin/streptomycin at 37 °C and 5% CO2, and split twice a week at a ratio of 1:3. The human non-seminomatous cell line NTERA-2 was originally obtained from ATCC and is reminiscent of an embryonal carcinoma [33 (link)]. The human seminoma-derived cell line TCam-2 was provided by Daniel Nettersheim (University of Düsseldorf, Germany) and previously used in our laboratory [20 (link)]. The initial passage number of both cell lines was 4, and another 5–10 passages were performed in the course of the experiments.
9
Establishing 4T1 Murine Mammary Carcinoma Model
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For both in vitro and in vivo studies, the 4T1 murine mammary carcinoma cell line, provided by Judy Lieberman (Lieberman Laboratory, Harvard University, Boston, MA, USA), was used. The 4T1 cells were grown as adherent culture in Dulbecco’s Modified Eagle Medium (DMEM high glucose, 4.5 g/L without l glutamine and Phenol Red, Capricorn Scientific, Ebsdorfergrund, Germany, Cat-No. DMEM-HXRXA) supplemented with 10% Fetal Bovine Serum (FBS—South America Origen, EU approved, EuroClone S.p.A., Pero, Italy, CatNo. ECS0180L), l -glutamine 200 mM (Capricorn Scientific, Ebsdorfergrund, Germany, Cat-No. GLN-B), and penicillin/streptomycin 100x (Capricorn Scientific, Ebsdorfergrund, Germany, Cat-No. PS-B). Cells were submitted to passages every 2 or 3 days. Trypsin 10x (Lonza A. G., Basel, Switzerland, Cat-No. 17-160E) was used to release cells from sub-confluent monolayers. The detached cells were seeded back into cell culture flasks or prepared for experiments. KRIBB11 was purchased from MedChemExpress (Monmouth Junction, NJ, USA, Cat-No. HY-100872).
10
Cell Culture Maintenance Protocol
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The cell lines used were human cancer cell lines: 518A2 (melanoma), A2780 (ovarian carcinoma), HT29 (colon adenocarcinoma), MCF-7 (breast adenocarcinoma), 8505C (thyroid carcinoma) and non-malignant mouse fibroblasts NIH 3T3. Cultures were maintained as monolayers in RPMI 1640 medium with l -glutamine (Capricorn Scientific GmbH, Ebsdorfergrund, Germany) supplemented with 10% heat inactivated fetal bovineserum (Sigma-Aldrich Chemie GmbH, Steinheim, Germany) and penicillin/streptomycin (Capricorn Scientific GmbH, Ebsdorfergrund, Germany) at 37 °C in a humidified atmosphere with 5% CO2.
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