Hello, I am working with mice that develop mammary tumors (PyMT transgenic mice) and I am trying to isolate epithelial cells from their tumors, however every time most of the cells that I isolate are just fibroblasts. Does anyone have suggestions as to how to get rid of them or what is the best protocol to use to increase the yield of epithelial cells? Thank you in advance.
HI Marta, See this article: http://mcr.aacrjournals.org/content/9/1/25.full.pdf+html
They have used Dynabeads Sheep anti-Rat IgG (Invitrogen by Thermo Fisher) and conjugated it to a rat anti mouse Thy-1 mAb for fibroblast depletion.
If available, you might consider FACS or MACS using EpCAM or any other epithelial specific marker.
In their paper, Díaz-Guerra and colleages describe a protocol for mammary gland digestion and they include a centrifugation step at 134 x g, 1 minute, to get rid of contaminating fibroblasts. Hopefully this also applies to mammary tumour digestion.
Article Intrinsic cues and hormones control mouse mammary epithelial tree size
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