Prior to MoFlo sorting, single cell suspensions were stained with the following antibodies: FITC-conjugated anti-CD11b (M1/70) (Miltenyi, Bergisch Gladbach, Germany), PerCP-conjugated anti-CD3ε (145-2C11) and PerCP-conjugated anti-CD19 (1D3) were purchased from BioLegend (San Diego, CA, USA), APC-conjugated anti-CD117 (c-kit) (2B8), PE-conjugated anti-CD27 (LG.3A11), PE-Cy7-conjugated anti-NK1.1 (PK136) and BD Horizon V450 conjugated anti-CD45.1 (A20) were purchased from BD Biosciences (Heidelberg, Germany). NK cell purity, functional properties and subset distribution was analyzed by FACS Canto II (BD Biosciences, Heidelberg, Germany) and use of the FlowJo Software (Treestar, Ashland, Oregon, USA).
Extracted molecule
total RNA
Extraction protocol
RNA was isolated from freshly sorted NK subsets using the RNAeasy Micro Kit (QIAGEN, Hilden, Germany) according to the manufacturer’s instructions. RNA quality was checked using the Agilent 2100 Bioanalyzer.
Label
Cy3
Label protocol
High-quality RNA were amplified and Cyanine 3-CTP-labelled with the one colour low RNA input linear amplification kit (Agilent).
Hybridization protocol
Labeling efficiency was controlled using the NanoDrop spectrophotometer and labeled cRNA were fragmented and hybridized on whole mouse genome expression arrays (4 × 44 K Agilent). Microarrays were washed and subsequently scanned with an Agilent scanner.
Scan protocol
Microarrays were washed and subsequently scanned with an Agilent scanner.
Description
SAMPLE 3
Data processing
Raw data were extracted with Feature Extraction 9.5.1 software and analyzed using GeneSpring GX 11 (Agilent).
