|
| Status |
Public on Mar 07, 2014 |
| Title |
Naïve CD4.Th17.TMP776.60h #1 |
| Sample type |
RNA |
| |
|
| Source name |
CD4+CD45RA+
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: CD4+
treatment: skewed under Th17 conditions plus plus TMP776 for 60 h
|
| Treatment protocol |
RORgt inhibitor (200 nM) or vehicle control (0.1% v/v DMSO) were added to the T cells in culture immediately before the addition of anti-CD3/anti-CD28 beads for diffferent time point.
|
| Growth protocol |
Human CD4+ T cells were cultured in standard T cell culture medium (10% FBS IMDM, 2 mM L-glutamine, 1 mM Pyruvate,0.1 mM NEAA, 0.05 mM 2-mercaptoethonol, 10 U/10 mg Penicillin/Streptomycin) at a density of 1 x1 0^6 per mL.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using the Qiagen RNeasy with on-column DNase treatment method.
|
| Label |
biotin
|
| Label protocol |
biotin
|
| |
|
| Hybridization protocol |
The labeled fragmented DNA was hybridized to the Gene Arrays 1.0ST for 16-18 hours in GeneChip Hybridization oven 640 at 45 °C with rotation (60 rpm). The hybridized samples were washed and stained using Affymetrix fluidics station 450. The first stain with streptavidin-R-phycoerythrin (SAPE) was followed by signal amplification using a biotinilated goat anti-streptavidin antibody and another SAPE staining (Hybridization, Washing and Staining Kit, Affymetrix).
|
| Scan protocol |
Standard Affymetrix protocol on an Affymetrix GeneArray Scanner 3000 7G Plus (Affymetrix).
|
| Description |
Naïve CD4+ T cells, purified by Naïve CD4+ T cells Isolation Kit II (human) from Miltenyi Biotec (cat No.: 130-094-131), skewed under Th17 conditions (Cytokine and Ab Cocktail (Final concentrations): IL-1b(10 ng/ml); IL-6 (20 ng/ml), IL-23 (10 ng/ml); TGFb (2 ng/ml); IL-2 (1ng/ml), a IL-4 (2 ug/ml); a IFNg (2 ug/ml)) plus TMP776 for 60 h
|
| Data processing |
Samples were RMA normalized using Affymetrix Power Tools (gcbg background correction, quantile normalization, median polish) on the annotated set of probesets. Unannotated probesets were removed prior to normalization (see supplemental .mps file)
|
| |
|
| Submission date |
Dec 18, 2013 |
| Last update date |
Sep 01, 2016 |
| Contact name |
Scott Davis |
| Organization name |
GSK
|
| Department |
Tempero DPU
|
| Street address |
200 Technology Square, Suite 602
|
| City |
Cambridge |
| State/province |
MA |
| ZIP/Postal code |
02139 |
| Country |
USA |
| |
|
| Platform ID |
GPL6244 |
| Series (1) |
| GSE53455 |
Expression profiling comparisons of human CD4+ T cells treated with RORgt inhibitors |
|
| Relations |
| Reanalyzed by |
GSE86357 |
