Fluorescent protein-assisted purification for gene expression profiling.
Cell type-specific expression of fluorescent proteins allows the purification of rare cells from complex -tissues by flow cytometry. This strategy is especially useful for molecular analysis of cancer cells because these cells can be effectively purified away from the noncancerous tumor stroma. Coexpression of bioluminescence with fluorescence makes further allows in vivo tracking of cancer cells, which can then be purified at specific tumorigenic stages. Here, we describe protocols for purifying rare skin stem cells, and for in vivo monitoring and purification of cancer cells from lung metastases. Also described is a protocol for the isolation of total RNA from the purified cells for the purpose of performing gene expression profiling.
Laboratory of Cancer Biology and Genetics, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD, USA.
M Raza
MR
Zaidi
Chi-Ping
CP
Day
Glenn
G
Merlino
D000818:Animals; D002469:Cell Separation / Q000379:methods*; D058948:Cell Tracking; D005434:Flow Cytometry; D020869:Gene Expression Profiling / Q000379:methods*; D005786:Gene Expression Regulation; D018014:Gene Transfer Techniques; D005822:Genetic Vectors / Q000235:genetics / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D008175:Lung Neoplasms / Q000235:genetics / Q000378:metabolism / Q000556:secondary; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D012867:Skin / Q000166:cytology; D013234:Stem Cells / Q000378:metabolism
D049452:Green Fluorescent Proteins