title: Th1 rep2
organism: Mus musculus
description: Protocols: CD4+, CD62L+ cells were purified with CD4+CD62L+ T Cell Isolation Kit II (Miltenyi Biotec). Cells were seeded into anti-CD3 (2 ug/ml, clone 145-2C11, eBioscience) and anti-CD28 (5 ug/ml, clone 37.51, eBioscience) coated 96-well plates, at a density of 250,000-500,000 cells/ml and cultured in a total volume of 200 ul in the presence of recombinant murine IL-12 (10 ng/ml, RD Systems) and neutralising anti-IL4 (10 ug/ml, clone 11B11, eBioscience). Cells were cultured for another four days in absence of CD3 and CD28 stimulation to rest them. The original medium was kept and fresh medium containing the same cytokines as before was added to dilute the cultures 1:3. Finally, the cells were labelled with anti-CXCR3-APC (1:200, clone CXCR3-173, BioLegend) and Propidium Iodide and FITC-conjugated antibodies against CD11b, CD11c, Ly6G, CD8a and CD19. The cultures were then FACS-sorted to obtain Propidium Iodide-, FITC-negative and CXCR3-positive cells. RNA was purified using the Qiagen RNEasy Plus Mini Kit automated using a QIAcube Samples were processed using the TruSeq RNA Sample Prep v2 kit (Illumina) according to the manufacturers instructions with an alteration to use KAPA Hifi polymerase for PCR amplification instead of the one supplied with the kit.
attributes.tag: ENA-FIRST-PUBLIC
attributes.value: 2015-04-05T17:04:12Z
attributes.tag: ENA-LAST-UPDATE
attributes.value: 2018-03-08T17:48:54Z
attributes.tag: External Id
attributes.value: SAMEA2536561
attributes.tag: INSDC center name
attributes.value: EBI
attributes.tag: INSDC first public
attributes.value: 2015-04-05T17:04:12Z
attributes.tag: INSDC last update
attributes.value: 2018-03-08T17:48:54Z
attributes.tag: INSDC status
attributes.value: public
attributes.tag: Submitter Id
attributes.value: E-MTAB-2582:Th1 rep2
attributes.tag: broker name
attributes.value: ArrayExpress
attributes.tag: cell type
attributes.value: T-helper 1 cell
attributes.tag: common name
attributes.value: house mouse
attributes.tag: sample name
attributes.value: E-MTAB-2582:Th1 rep2
attributes.tag: scientific_name
attributes.value: Mus musculus