characteristics.text: ERC000011
characteristics.characteristic: ENA-CHECKLIST
characteristics.text: 2013-11-01T17:01:38Z
characteristics.characteristic: ENA-FIRST-PUBLIC
characteristics.text: 2018-03-08T16:48:41Z
characteristics.characteristic: ENA-LAST-UPDATE
characteristics.text: SAMEA2224113
characteristics.characteristic: External Id
characteristics.text: Department of Experimental Immunology, Academic Medical Center of the University of Amterdam
characteristics.characteristic: INSDC center name
characteristics.text: 2013-11-01T17:01:38Z
characteristics.characteristic: INSDC first public
characteristics.text: 2018-03-08T16:48:41Z
characteristics.characteristic: INSDC last update
characteristics.text: public
characteristics.characteristic: INSDC status
characteristics.text: ERS360462
characteristics.characteristic: SRA accession
characteristics.text: E-MTAB-1969:SAMPLE2_MACROPHAGES
characteristics.characteristic: Submitter Id
characteristics.text: ArrayExpress
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0009737
characteristics.characteristic: broker name
characteristics.text: monocyte
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/CL_0000576
characteristics.characteristic: cell type
characteristics.text: human
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/NCBITaxon_9606
characteristics.characteristic: common name
characteristics.text: Protocols: Peripheral blood mononuclear cells (PBMCs) were isolated from buffy coats using Lymphoprep density gradient centrifugation. Monocytes were isolated by adherence to plastic and cultured in Iscove's modified Dulbecco's medium supplemented with 10% [v/v] heat-inactivated human pooled serum (HPS), penicillin (100 U/ml), streptomycin (100 ug/ml) and ciproxin (5 ug/ml) at 37C in a humidified atmosphere supplemented with 5% CO2. Monocytes were incubated for 5 days in the presence of different cytokines: IFN-gamma (250 U/ml) in combination with TNF-alpha (12.5 ng/ml) (M1), IL-4 (50 ng/ml) (M2a), IL-10 (50 ng/ml) (M2c) or medium alone. Total RNA was isolated from monocytes and 5-day cultured macrophages (unstimulated or cytokine-polarized), using TriPure Isolation Reagent (Roche) according to the manufacturers instructions. Library preparation was carried out following the SOLiD Total RNA-Seq Kit Protocol (PN 4445374, Life Technologies) according to the manufacturers protocol for Small RNA library preparation. Briefly, RNA samples were first enriched for their small RNA fraction, by size selection between 15nt and 40nt on 15% TBE-Urea acrylamide gels. The small RNA-enriched samples were hybridized and ligated for cDNA synthesis. cDNA libraries were further purified with the MinElute PCR Purification Kit (Qiagen). After a size-selection step on 10% TBE-Urea gels, cDNA samples were amplified on gel using 19 cycles instead of 15 recommended by the manufacturer. For multiplexing purposes, the cDNA samples were amplified using the SOLiD 3' Primers from the SOLiD RNA Barcoding Kit. cDNA libraries were purified using the PureLink PCR Micro Kit (Invitrogen) and subjected to a second round of size selection on gel and purification according to the manufacturers instructions. The quality of the cDNA library was assessed with a smear analysis using the Bioanalyzer 2100 software (Agilent).
characteristics.characteristic: description
characteristics.text: normal
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/PATO_0000461
characteristics.characteristic: disease
characteristics.text: Donor 1
characteristics.characteristic: individual
characteristics.text: Homo Sapiens
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/NCBITaxon_9606
characteristics.characteristic: organism
characteristics.text: Homo sapiens
characteristics.characteristic: organism
characteristics.text: Homo sapiens
characteristics.characteristic: scientific_name
characteristics.text: fresh specimen
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/OBI_0000971
characteristics.characteristic: specimen with known storage state
characteristics.text: SAMPLE2_MACROPHAGES
characteristics.characteristic: title