Oncosphere Assay for Cancer Cells

Oncosphere assays were performed as previously described30 (link). Briefly, cancer cells were collected by using scraper and filtered using 40 μm pore cell strainer (BD Biosciences). 0.5–2.5 × 10⁴ single cells were seeded into 6-well ultra-low binding plates (Corning) and cultured in Human mammary epithelial cell (HuMEC) media (Life Technologies) containing 20 ng ml⁻¹ EGF, 5 μg ml⁻¹ insulin, 10 μg ml⁻¹ heparin, 40 ng ml⁻¹ basic FGF, 1X B27 supplement without vitamin A and 0.02% methylcellulose for 7 days. Spheres over 100 μm diameter were counted and pictured using Nikon ECLIPSE TS100 microscope.

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Song K.H., Park M.S., Nandu T.S., Gadad S., Kim S.C, & Kim M.Y. (2016). GALNT14 promotes lung-specific breast cancer metastasis by modulating self-renewal and interaction with the lung microenvironment. Nature Communications, 7, 13796.

Publication 2016

40 μm pore cell strainer 6-well ultra-low binding plates HuMEC) media ECLIPSE TS100 microscope

Assays Cancer Cells Cultured media Epithelial cell Heparin Human Insulin Mammary Methylcellulose Microscope Supplement Vitamin a

Corresponding Organization : Korea Advanced Institute of Science and Technology

Other organizations : The University of Texas Southwestern Medical Center, Korea National Institute of Health

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Variable analysis

independent variables

Seeding density (0.5–2.5 × 10^4 single cells)

dependent variables

Number of spheres over 100 μm diameter

control variables

Cell culture conditions (6-well ultra-low binding plates, HuMEC media with 20 ng ml^-1 EGF, 5 μg ml^-1 insulin, 10 μg ml^-1 heparin, 40 ng ml^-1 basic FGF, 1X B27 supplement without vitamin A, 0.02% methylcellulose)
Culture duration (7 days)

controls

No positive or negative controls were explicitly mentioned.

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