characteristics.text: ERC000011
characteristics.characteristic: ENA-CHECKLIST
characteristics.text: 2018-01-17T17:05:16Z
characteristics.characteristic: ENA-FIRST-PUBLIC
characteristics.text: 2017-03-14T16:47:13Z
characteristics.characteristic: ENA-LAST-UPDATE
characteristics.text: SAMEA103910985
characteristics.characteristic: External Id
characteristics.text: University of Michigan School of Public Health
characteristics.characteristic: INSDC center name
characteristics.text: 2018-01-17T17:05:16Z
characteristics.characteristic: INSDC first public
characteristics.text: 2017-03-14T16:47:13Z
characteristics.characteristic: INSDC last update
characteristics.text: public
characteristics.characteristic: INSDC status
characteristics.text: ERS1600153
characteristics.characteristic: SRA accession
characteristics.text: E-MTAB-5585:Sample24_1_MET
characteristics.characteristic: Submitter Id
characteristics.text: ArrayExpress
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0009737
characteristics.characteristic: broker name
characteristics.text: ALDH+ cell
characteristics.characteristic: cell type
characteristics.text: human
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/NCBITaxon_9606
characteristics.characteristic: common name
characteristics.text: Protocols: Normal (non-pathogenic) breast tissue was isolated from women undergoing voluntary reduction mammoplasty at the University of Michigan. The study protocol was approved by the University of Michigan Institutional Review Board. Breast tissue was mechanically and enzymatically digested as previously described (Dontu et al., 2003; Kakarala et al., 2010). Normal mammary cells were stained for CD44, and CD24 expression and ALDH activity accessed by the Aldefluor assay as described previously (Colacino et al., 2016) and analyzed or sorted on a Beckman Coulter MoFlo Astrios at the University of Michigan Flow Cytomtery core facility. Briefly, single mammary cells were first incubated with a lineage depletion cocktail that consisted of biotinylated antibodies targeted against CD45, HLA-DR, CD14, CD31, CD41, CD19, CD235a, CD56, CD3, CD16, and CD140b (all from eBioscience, except for CD140b (Biolegend) and CD41 (Acris)). Next, cells were stained with Alexafluor 750 tagged streptavidin, LIVE/DEAD Fixable Dead Cell Stain (Invitrogen), CD24 (Biolegend), CD44 (BD), and Aldefluor (Stem Cell Technology). Single color and isotype controls were included for compensation and gating purposes. Aldefluor-positive gating was based on DEAB (negative) controls. Flow cytometry data analysis was performed with FlowJo software v10.0.8. Total RNA was isolated from each cell population using the RNEasy Micro Kit (Qiagen) with on column DNase treatment. RNA concentration and quality was determined using a Nanodrop (Thermo) and Bioanalyzer (Agilent). Ribosomal RNAs were depleted using Ribominus (Life) and sequencing libraries were prepared with the SMARTer Stranded RNA-Seq kit (Clontech).
characteristics.characteristic: description
characteristics.text: normal
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/PATO_0000461
characteristics.characteristic: disease
characteristics.text: 24
characteristics.characteristic: individual
characteristics.text: Homo sapiens
characteristics.characteristic: organism
characteristics.text: Homo sapiens
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/NCBITaxon_9606
characteristics.characteristic: organism
characteristics.text: mammary gland
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/BTO_0000817
characteristics.characteristic: organism part
characteristics.text: Homo sapiens
characteristics.characteristic: scientific_name
characteristics.text: Sample24_1_MET
characteristics.characteristic: title