Scid beige

All animals used in this study received humane care in compliance with the principles stated in the Guide for the Care and Use of Laboratory Animals, NIH Publication, 1996 edition, and the protocol was approved by the Institutional Animal Care Committee of Universityof Washington. The C57BL/6 and SCID/Beige mice were purchased from Charles River (Wilmington, MA). Lgr5-DTR-eGFP mice were originally generated by Genentech (Tian et al., 2011 (link)) and were kindly provided by Dr. Noah Shroyer at the Baylor College of Medicine. Krt7-CreER^T2 was kindly provided by Dr. Jianwen Que at Columbia University. Krt8-CreER^T2 was made by our laboratory as described (Zhang et al., 2012 ). C57BL/6-Gt(ROSA)26Sor^{tm1(HBEGF)Awai}/J, B6.Cg-Gt(ROSA)26Sor^{tm14(CAG-tdTomato)Hze}/J, B6.129P2-Lgr5^{tm1(cre/ERT2)Cle}/J, Col1a2-CreER^T2, Pdgfrβ-CreER^T2, NG2-CreER^T2, B6.Cg-Gt(ROSA)26Sor^{tm3(CAG-EYFP)Hze}/J mice were from the Jackson Laboratory (Bar Harbor, ME). Male mice at the age of E15.5 to postnatal 27 weeks were used. Mice were genotyped by polymerase chain reaction using mouse genomic DNA from tail biopsy specimens. The sequences of genotyping primers and the expected band sizes for PCR are listed in Table S3. PCR products were separated electrophoretically on 1% agarose gels and visualized via ethidium bromide under UV light.

Animals were housed and handled in accordance with protocols approved by the NCI IACUC. 8-10 week old female BALB/c mice (Charles River) were used for allograft experiments. 8-10 week old female SCID-beige (Charles River) mice were used for xenograft experiments. Pharmacokinetic studies were completed in 8-10 week old female Athymic Nude mice (Charles River).

2.5×10⁶ (Figure 1, S1), 5×10⁶ (Figure 4, S3) or 1×10⁷ (Figure 6) cells were resuspended in phosphate buffered saline (PBS), mixed 1:1 with Matrigel and injected subcutaneously into the flanks of SCID/beige (Charles River Laboratory), or Athymic Nude-FoxN1^nu mice (Harlan). Tumor volumes were determined twice per week and calculated as (length × width²)π/6. Mice were sacrificed when the tumors reached 1000mm³ or the mice exhibited signs of moribundity, at which point tumors were removed and weighed. Tumors were halved at harvest and formalin-fixed paraffin-embedded or flash frozen. Experiments were approved by the Duke University Institutional Animal Care and Use Committee and the University of Virginia Animal Care and Use Committee.

All animal experiments were performed at the UAB under an Institutional Animal Care and Use Committee (IACUC)-approved protocol (#20290) in accordance with NIH guidelines. Immunocompromised mice (SCID Beige) were purchased from Charles River. For intracardiac injection, mice were anesthetized with ketamine/xylazine and fixed in place. The needle was inserted into the left ventricle of the heart properly from the under-costal space. When blood was observed pumping into the needle tube, we began to slowly inject the cell solution. Mice were placed on the stage warmed at 37°C until they were fully awake. For intracranial injection, dissociated breast cancer cells were stereotactically injected into the striatum of mice as described (28 (link)).