|
Status |
Public on Apr 11, 2010 |
Title |
Cord blood CD4+_act+IL-12_48h_rep1 |
Sample type |
RNA |
|
|
Source name |
human cord blood CD4+ T cell, activated and IL-12 treated, 48h
|
Organism |
Homo sapiens |
Characteristics |
cell source: human neonatal umbilical cord blood cell type: CD4+ T cell
|
Treatment protocol |
Cells were cultured on 24-well plates, 2x106 cells/1ml Yssel´s medium, activated via T cell receptor (plate bound antiCD3 500 ng/well, and soluble antiCD28 500 ng/ml, Immunotech, France). Cytokines IL-12 (2.5 ng/ml, R&D Systems), highly purified human leukocyte IFN-alpha (100 U/ml, Red Cross Finland Blood Service) or the combination of IL-12 (2.5 ng/ml) + IL-18 (25 ng/ml, Medical & Biological Laboratories) were added to induce Th1 differentiation. Cells were harvested at 2h, 6h or 48h.
|
Growth protocol |
Umbilical cord blood was collected from healthy neonates born in Turku University Hospital, Hospital District of Southwest Finland. Mononuclear cells were isolated with Ficoll-Paque (Amersham Biosciences) gradient centrifugation, and CD4+ cell population was purified using magnetic beads (Dynal CD4 Positive Isolation Kit, Invitrogen) according to manufacturers instructions.
|
Extracted molecule |
total RNA |
Extraction protocol |
Total RNA was extracted with TRIzol reagent (Invitrogen) and further purified with Rneasy mini kit (Qiagen) according to the manufacturer's instructions. RNA was eluted in dH2O.
|
Label |
biotin
|
Label protocol |
Amplification was started from 5 micrograms of total RNA. The cDNA/cRNA synthesis reactions and sample cleanup steps were performed according to Affymetrix´s GeneChip Expression Analysis Technical Manual.
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|
|
Hybridization protocol |
The biotinylated and fragmented cRNA was hybridized to Affymetrix GeneChip Human Genome U133A arrays overnight (16-18 hours) at 45C. GeneChips were washed and stained in the Affymetrix Fluidics Station.
|
Scan protocol |
GeneChips were scanned using the GeneChip Scanner 3000 with AutoLoader.
|
Description |
biological replicate 1
|
Data processing |
The data was analyzed using MAS 5.0. Data was normalized to target intensity 50.
|
|
|
Submission date |
Feb 04, 2010 |
Last update date |
Sep 01, 2016 |
Contact name |
Sanna Filén |
Organization name |
Turku Centre for Biotechnology
|
Street address |
Tykistökatu 6
|
City |
Turku |
ZIP/Postal code |
20520 |
Country |
Finland |
|
|
Platform ID |
GPL96 |
Series (1) |
GSE20198 |
ATF3 is a positive regulator of human IFN gene expression |
|
Relations |
Reanalyzed by |
GSE86363 |