DMEM/F-12

Protocol tips

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	Seed 4 × 104 cells/ml to an ultra-low attachment tray

Protocol tips
Seed 4 × 104 cells/ml to an ultra-low attachment tray

Publication protocol

Mouse mammary glands/tumors were dissected, with removal of the lymph node if possible. Tissue was manually dissociated and then digested in 10% collagenase/hyaluronidase (Stem Cell Technologies, Vancouver, BC, Canada) in Dulbecco's Modified Eagle's medium (DMEM) for 3-4 h with gentle tilting. Single cells were extracted and filtered through a 70 μm nylon mesh. To remove contaminating infiltrating cells of hematopoietic origin, Biotin Binder Dynabeads (Invitrogen) in combination with a biotinylated anti-mouse lineage antibody panel (BioLegend, San Diego, CA, USA) were used as suggested by the manufacturer. For mammosphere assay, cells were seeded in mammosphere medium (1:1 mixture of DMEM and Ham’s F12 medium (Gibco, Life Technologies, Mulgrave, VIC, Australia) supplemented with 1 × B27 (Invitrogen), 20ng/ml epidermal growth factor, 20 ng/ml basic fibroblast growth factor, 4 μg/ml heparin (Sigma-Aldrich, St Louis, MO, USA), penicillin–streptomycin, and 0.25 μg/ml fungazone) at 4 × 104 cells/ml to an ultra-low attachment tray (Corning Inc., Corning, NY, USA).

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Manufacturer protocol

Download the product protocol from Thermo Fisher Scientific for DMEM/F-12 below.

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