BT4Ca (Institute of Cell Biology, Department of Cancer Research, University of Essen Medical School, Germany), C6, F98 and RG-2 cells (Uniklinikum Erlangen, Neuro-oncological Research Laboratory) were cultured in tissue culture dishes (Sarstedt AG & Co, Nümbrecht, Germany) in Dulbecco’s modified Eagle’s medium (DMEM, FG 0445, Biochrom GmbH, Berlin, Germany), supplemented with 10% heat-inactivated foetal calf serum (Biochrom GmbH), 1 mg/ml penicillin and 0.1 mg/ml streptomycin (Biochrom GmbH) and 1 x non-essential amino acids (Biochrom GmbH). Cells were kept in an incubator at 37 °C with 5% CO2 and split once to twice a week at 80–90% confluence.
Tissue culture dishes
Manufactured by Sarstedt
Sourced in Germany
Tissue culture dishes are flat, circular containers used for the growth and maintenance of cells, tissues, or organisms in a controlled environment. They provide a sterile surface for culturing and observing biological samples.
Automatically generated - may contain errors
Lab products found in correlation
Glutamax, by Thermo Fisher Scientific (3 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (3 mentions)
Turbofect, by Thermo Fisher Scientific (2 mentions)
Foetal calf serum, by Harvard Bioscience (1 mentions)
Penicillin, by Harvard Bioscience (1 mentions)
Streptomycin, by Harvard Bioscience (1 mentions)
Non essential amino acids (neaa), by Harvard Bioscience (1 mentions)
Fg 0445, by Harvard Bioscience (1 mentions)
Tcs spe, by Leica (1 mentions)
Prolong gold antifade, by Merck Group (1 mentions)
Trypsin, by BD (1 mentions)
Sodium pyruvate, by Fujifilm (1 mentions)
Ethylene diamine tetraacetic acid (edta), by Fujifilm (1 mentions)
Hek293t cells, by RIKEN Cell Bank (1 mentions)
Recombinant human basic fgf, by Fujifilm (1 mentions)
Ultra low attachment surface culture dishes, by Corning (1 mentions)
Recombinant human egf, by Fujifilm (1 mentions)
Dmem f12, by Fujifilm (1 mentions)
B 27 supplement without vitamin a, by Thermo Fisher Scientific (1 mentions)
Penicillin streptomycin, by Fujifilm (1 mentions)
8 protocols using tissue culture dishes
1
Culturing Glioma Cell Lines
2
Appressorium Formation and Penetration Assay
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Tissue culture dishes (60 x 15 mm; Sarstedt AG; Germany) were coated with 1,16-hexadecanediol (1 ml solution, 0.1 mM in methanol), which is inductive for appressorium formation in M. grisea [32 (link)]. Though not essential, this led to more reliable results across different charges of plastic plates. After drying, 5 ml of conidia suspension (2 x 105 conidia/ml) were added and incubated for 24 h at 25°C. To check the ability to penetrate a cellophane surface, sterilized dialysis tubes (Visking 36/32, Carl Roth, Karlsruhe, Germany) were put on 1.2% agarose-covered glass slides and incubated with 1 x 105 conidia for two days at 25°C and high humidity conditions. The incipient cytorrhysis assay of appressoria was performed in coated Tissue culture dishes with 1 x 106 conidia/dish. The water was substituted after 24 h with polyethylene glycol 6000 solutions with concentrations varying from 100 to 500 mg/ml which corresponds to external turgor pressures from 70 Pa to 5.8 MPa [33 (link)]. The percentage of collapsed appressoria was quantified after 10 min of incubation in polyethylene glycol solution in three biological replicates with at least 100 appressoria each.
3
Subcellular Localization of DdTMEM16-GFP
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HEK293 cells were seeded on glass coverslips in tissue culture dishes 35×10 mm (Sarstedt, 83.1800) and grown to 90% confluence for 48 h in DMEM supplemented with GlutaMax (Life Technologies), 10% fetal calf serum and 1% penicillin/streptomycin (PAA). Confluent cells were transfected with Turbofect (Thermo Scientific) according to the manufacturer’s instructions. After 24 h of protein expression, cells were washed with PBS buffer and fixed with 4% PFA for 15 min at room temperature. Afterwards, cells were washed with PBS buffer and mounted on microscopy slides with ProLong Gold Antifade (Sigma, 36930). Localization of DdTMEM16-GFP protein in HEK293 cells was analyzed by imaging fluorescence with a Leica SPE confocal microscope (Leica, TCS SPE).
4
Culturing and Differentiating Osteosarcoma Stem Cells
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HEK293T cells were obtained from the RIKEN Cell Bank (Saitama, Japan) and cultured in DMEM (FUJIFILM Wako Pure Chemical) supplemented with 10% FBS (Hyclone) and 1% penicillin/streptomycin (Thermo Fisher Scientific) at 37°C in 5% CO2 (42 (link)). The patient-derived OS cell line 143B was obtained from the ATCC (Manassas, USA) and cultured in adherent medium containing DMEM supplemented with 10% FBS, 110 μg/mL sodium pyruvate (FUJIFILM Wako Pure Chemical), and 1% penicillin/streptomycin. Both cell types were cultured in tissue culture dishes (SARSTEDT) to ensure optimal adherence and expansion. To enrich stem-like cells, 143B cells were harvested using trypsin (BD Bioscience) and EDTA (FUJIFILM Wako Pure Chemical), then cultured in osteosphere medium containing DMEM/F12 (FUJIFILM Wako Pure Chemical) supplemented with 20 ng/mL recombinant human EGF (FUJIFILM Wako Pure Chemical), 20 ng/mL recombinant human basic FGF (FUJIFILM Wako Pure Chemical), B27 supplement without vitamin A (Gibco), GlutaMAX (Thermo Fisher Scientific), and 1% penicillin/streptomycin. Under these conditions, the cells were incubated in Ultra-Low Attachment Surface culture dishes (Corning). To assess the differentiation potential of OSCs, the cells were transferred from osteosphere to adherent medium, and from Ultra-Low Attachment Surface to tissue culture dishes, to promote adherence and differentiation.
5
Microelectrode Fabrication and Calibration
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Microelectrodes were made by pulling (pipette puller PIP 6, HEKA, Lambrecht, Germany) from glass capillaries (Ø1.5 mm, Hilgenberg, Malsfeld, Germany), baked at 210 °C (LE040K1RN, Nabertherm, Lilienthal, Germany), and silanized (Tributylchlorosilane, Fluka, Buchs, Switzerland) for achieving appropriate shape and hydrophobic surface. The tip diameter was adjusted to approximately 10 μm. As the inner (backfilling) solution, 200 mM KCl was used. The electrode tip was filled with a liquid ion exchanger (LIX) (potassium ionophore I—Cocktail A, FLUKA: for K+-flux measurements). As a reference electrode, a pipette with tip diameter of about 100 µm filed with 1 M NaCl in 2% agarose was used. A chlorided silver wire was inserted in both electrodes and connected with the MIFE preamplifier.
Before each experiment, calibration was carried out (six calibration points in the range between 10 μM and 3 mM of the respective ion). Calibrations were performed in tissue culture dishes (Ø35 mm, Sarstedt, Nümbrecht, Deutschland) filled with 2 mL of calibration solution (measurement solution without KCl).
Before each experiment, calibration was carried out (six calibration points in the range between 10 μM and 3 mM of the respective ion). Calibrations were performed in tissue culture dishes (Ø35 mm, Sarstedt, Nümbrecht, Deutschland) filled with 2 mL of calibration solution (measurement solution without KCl).
6
DMEM-based Cell Culture Protocol
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Dulbecco’s Modified Eagle’s Medium (DMEM), fetal calf serum (FCS), methionine-free DMEM, streptomycin, penicillin, protease inhibitors, trypsin, trypsin-EDTA, DEAE-dextran, protein A-sepharose, and triton X-100 were purchased from Sigma-Aldrich. Tissue culture dishes were purchased from Sarstedt (Nuembrecht, Germany). [35S]-methionine was obtained from PerkinElmer (Waltham, Massachusetts). Acrylamide, Tris, TEMED, SDS, dithiothreitol (DTT), paraformaldehyde, polyvinyl difluoride (PVDF) membrane, ProLong Gold Antifade mountant with DAPI by Invitrogen, and sucrose were purchased from Carl Roth GmbH (Karlsruhe, Germany). Lubrol WX was obtained from MP Biomedicals (Eschwege, Germany). Endo-β-N-acetylglucosaminidase H (endo H) was acquired from Roche Diagnostics (Mannheim, Germany). TRIzol reagent (Life Technologies, California, USA), restriction enzymes, Isis proofreading DNA polymerase, molecular weight standards for SDS-PAGE, and SuperSignal™ West Femto maximum sensitivity western blot chemiluminescence substrate were purchased from Thermo Fisher (Schwerte, Germany).
7
Chick Spinal Cord Explant Culture
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All animal experiments were carried out in accordance with the Canadian Council on Animal Care guidelines and approved by the IRCM Animal Care Committee and the McGill University Animal Care Committee. Fertilised chicken eggs (FERME GMS, Saint-Liboire, QC, Canada) were incubated (Lyon Technologies, model PRFWD) at 39 °C according to standard protocols25 (link). LMC explants were collected from HH st. 24–26 lumbar spinal cords and incubated in 95% air and 5% CO2 at 37 °C in MN medium for about 18 hours as previously described27 (link). 20 mL of MN medium solution is: 19.2 mL of Neurobasal medium (Invitrogen, cat. no. 21103-049), 400 µL Serum-free supplement (50×, B-27; Invitrogen, cat. no. 17504-044), 2 µL of l-glutamic acid (50 mM, Sigma-Aldrich, cat. no. G8415), 73 mg of l-glutamine (Invitrogen, cat. no. 21051-024) and 200 µL of penicillin-streptomycin (100×, Invitrogen, cat. no. 15140-122). Prior to explant culture, tissue culture dishes (Sarstedt, cat. no. 83.3901.300) were coated with 20 µg/mL Laminin (Invitrogen, cat. no. 23017-015) for 2 hours at 37 °C and rinsed with Neurobasal medium.
8
pH Measurement of CAP-Exposed Solutions
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For pH measurement, 7 × 20 µL drops of the respective solution in 35 mm diameter tissue culture dishes (Sarstedt, Nümbrecht, Germany) were exposed to CAP and then transferred into a 1.5 mL reaction vessel. The pH measurement was performed immediately by using the LAQUAtwin pH meter (HORIBA Scientific, Kyōto, Japan).
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