title: Mitochondrial staining allows robust elimination of apoptotic and damaged cells during cell sorting.
abstract: High-speed fluorescence-activated cell sorting is relevant for a plethora of applications, such as PCR-based techniques, microarrays, cloning, and propagation of selected cell populations. We suggest a simple cell-sorting technique to eliminate early and late apoptotic and necrotic cells, with good signal-to-noise ratio and a high-purity yield. The mitochondrial potential dye, TMRE (tetramethylrhodamine ethyl ester perchlorate), was used to separate viable and non-apoptotic cells from the cell sorting samples. TMRE staining is reversible and does not affect cell proliferation and viability. Sorted TMRE(+) cells contained a negligible percentage of apoptotic and damaged cells and had a higher proliferative potential as compared with their counterpart cells, sorted on the basis of staining with DNA viability dye. This novel sorting technique using TMRE does not interfere with subsequent functional assays and is a method of choice for the enrichment of functionally active, unbiased cell populations.
authors.affiliation: Program in Cellular and Molecular Medicine, Children's Hospital Boston and Department of Pediatrics, Harvard Medical School, Boston, Massachusetts (NSB).
authors.forename: Natasha S
authors.initials: NS
authors.lastname: Barteneva
authors.forename: Eugeny D
authors.initials: ED
authors.lastname: Ponomarev
authors.forename: Alla
authors.initials: A
authors.lastname: Tsytsykova
authors.forename: Myriam
authors.initials: M
authors.lastname: Armant
authors.forename: Ivan A
authors.initials: IA
authors.lastname: Vorobjev
mesh_terms: D000818:Animals; D017209:Apoptosis; D002460:Cell Line; D049109:Cell Proliferation; D005434:Flow Cytometry / Q000379:methods*; D005456:Fluorescent Dyes; D006801:Humans; D008264:Macrophages / Q000166:cytology / Q000378:metabolism; D051379:Mice; D008928:Mitochondria / Q000378:metabolism*; D009942:Organometallic Compounds; D010587:Phagocytosis; D013194:Staining and Labeling
chemical_list: D005456:Fluorescent Dyes; D009942:Organometallic Compounds; C110932:tetramethyl rhodamine ethyl ester
keywords: Click-IT; TMRE; apoptotic cell elimination; caspase; cell sorting; mitochondrial dye; viability