For frozen sections, mammary tumors

For frozen sections, mammary tumors or lungs were fixed in formalin for 1 hour, cryoprotected in 30% sucrose with shaking overnight, and embedded in OCT. Tissue sections were washed 3 x 5 minutes in wash buffer (PBS-T containing 0.3% TritonX-100), blocked in 10% goat serum or 5% donkey serum for 1 hour, incubated with the primary antibodies for 8-12 hours at 4C in blocking buffer, washed 4 x 20 minutes in wash buffer, incubated with the secondary antibodies for 1-2 hours at room temperature in blocking buffer, washed 4 x 20 minutes with wash buffer, and mounted (Vectashield, Vector Labs). For paraffin sections, mammary tumors were fixed in formalin overnight, stored in 70% ethanol, and processed for paraffin embedding and sectioning. Tissue sections were de-paraffinized, rehydrated, and treated with sodium citrate buffer for antigen retrieval, before being stained in the same process as with frozen sections. Tumorspheres and dissociated single cells were processed for immunofluorescence by fixing them in a Matrigel bed with formalin for 30 minutes, then directly starting the immunostain process as with frozen sections. Primary antibodies