To evaluate the formation of primary spheres, Mz-ChA-1 and Sk-ChA-1 cells were transferred to ultralow-adherence six-well plates (Corning, Inc., Corning, NY, USA) at a density of 5000 per well in 4 ml of CSC medium consisting of DMEM/F12 plus glutaMAX, B27 supplement, 20 ng ml−1 epidermal growth factor and 20 ng l−1 basic fibroblast growth factor. Tumour spheres were allowed to grow for 7 days. To establish spheres, cells were individualised by enzymatic dissociation, strained, and then plated at a density of 5 × 102 or 1 × 103 cells per well in 24-well plates (3 wells per treatment). Cells were treated with rapamycin1 μmol l−1, vismodegib10 μmol l−1, or both for 7 days, after which the culture medium was changed to fresh CSC medium. The resultant spheres were photographed after 14 days of culture.
Ultralow adherence six well plates
Manufactured by Corning
Sourced in United States
Corning's Ultralow Adherence Six Well Plates are designed for culturing cells that tend to aggregate or detach from traditional cell culture surfaces. These plates feature a specialized surface treatment that minimizes cell attachment, promoting the formation of cell spheroids, organoids, or suspension cultures.
Automatically generated - may contain errors
Lab products found in correlation
Epidermal growth factor (egf), by Merck Group (3 mentions)
Cs fcs, by PAN Biotech (3 mentions)
B27 supplement, by Thermo Fisher Scientific (3 mentions)
Hydrocortisone, by Merck Group (3 mentions)
Insulin, by Merck Group (2 mentions)
Penicillin streptomycin, by Merck Group (2 mentions)
Dmem f12, by Thermo Fisher Scientific (1 mentions)
Capmatinib, by Selleck Chemicals (1 mentions)
Foretinib, by Selleck Chemicals (1 mentions)
Crizotinib, by Selleck Chemicals (1 mentions)
Capmatinib incb28060, by Selleck Chemicals (1 mentions)
5 protocols using ultralow adherence six well plates
1
Tumor Sphere Formation and Modulation
2
Generation of Mesenchymal Breast Cancer Cells
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Mesenchymal transformed MCF-7 breast cancer cells (MCF-7-EMT) were generated as described earlier (20 (link)). Briefly, 4 × 104 cells/ml were cultured in prolonged mammosphere culture (5–6 weeks) in ultralow adherence six well plates (Corning, Lowell, MA, USA) in DMEM/F12 (Gibco) supplemented with 10% charcoal-stripped fetal calf serum (cs-FCS;PAN-biotech, Aiden Bach, Germany), 2% B27 supplement (Invitrogen, Darmstadt, Germany), 1% penicillin/streptomycin, 0.5 mg/ml hydrocortisone (Sigma, St. Louis, MO, USA), 5 μg/ml insulin, 20 ng/ml epidermal growth factor (EGF; Sigma, St. Louis, MO, USA).
3
Assessing Melanoma Cell Viability
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MM370, MM455, and MM603 melanoma cells (5 × 105), with or without induction of BRN2 or lacZ with 50 ng/ml doxycycline, were plated into ultra-low-adherence six-well plates (Corning, Lowell, MA). Viable cells were counted after 7 days using trypan blue exclusion, and expressed as a percentage of total cells counted per hemocytometer grid. Crizotinib (PF-02341066), foretinib (GSK1363089), and capmatinib (INCB28060) (SelleckChem) were dissolved in DMSO and used at a final concentration of 0.25 µm (Crizotinib and foretinib) or 1.25 nm (capmatinib).
4
Mesenchymal Transformation of MCF-7 Breast Cancer Cells
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Mesenchymal transformed MCF-7 breast cancer cells (MCF-7-EMT) were generated as described earlier14 (link). Briefly, 4 × 104 cells/ml were cultured in prolonged mammosphere culture (5–6 weeks) in ultralow adherence six well plates (Corning, Lowell, MA, USA) in DMEM/F12 supplemented with 10% charcoal-stripped fetal calf serum (cs-FCS;PAN-biotech, Aidenbach, Germany), 2% B27 supplement (Invitrogen, Darmstadt, Germany), 1% penicillin/streptomycin, 0.5 mg/ml hydrocortisone (Sigma, St. Louis, MO, USA ), 5 µg/ml insulin, 20 ng/ml epidermal growth factor (EGF; Sigma, St. Louis, MO, USA).
5
Generation of Mesenchymal-Transformed Breast Cancer Cells
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Mesenchymal-transformed MCF-7 breast cancer cells (MCF-7-EMT) and mesenchymal-transformed T-47D breast cancer cells were generated as described earlier [22 (link)]. Briefly, 4 × 104 cells/mL were cultured in a prolonged mammosphere culture (5–6 weeks) in ultralow adherence six-well plates (Corning, Lowell, MA, USA) in DMEM/F12 supplemented with 10% charcoal-stripped fetal calf serum (cs-FCS; PAN-biotech, Aidenbach, Germany), 2% B27 supplement (Invitrogen, Darmstadt, Germany), 1% P/S, 0.5 mg/mL hydrocortisone (Sigma), 5 µg/mL insulin, and 20 ng/mL epidermal growth factor (EGF; Sigma).
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