|
| Status |
Public on Oct 21, 2008 |
| Title |
Infected_dendritic_cells_sample_3 |
| Sample type |
RNA |
| |
|
| Source name |
Dendritic cells infected with L.monocytogenes
|
| Organism |
Homo sapiens |
| Characteristics |
Inhibitory monocyte-derived human dendritic cells infected with L.monocytogenes.
|
| Treatment protocol |
After harvesting (day 7 of culture) immature DC were infected with Listeria monocytogenes (L.monocytogenes) at MOI of 10 and consequently incubated for 6 hours.
|
| Growth protocol |
Circulating CD14+ monocytes were positively enriched from PBMC, obtained from buffy coats of healthy donors using CD14 MicroBeads (cell purity > 95%). Immature monocyte-derived dendritic cells (immDC) were generated from the CD14+ monocytes in a serum-free medium (CellGro) supplemented with 2mM glutamine, 500 U/ml IL-4 and 800 U/ml GM-CSF for seven days (cytokines were replenished on day 3 and 5).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Monocyte-derived DC were harvested 6 hours after infection with L.monocytogenes, lyzed in TRIzol and stored at –80° until further processing. RNA was extracted from cell lyzates with chloroform/isopropanol method, and was clean-uped in silica columns (Qiagen). The quality of the isolated RNA was determined by measuring the A260 / 280 ratio and the integrity of the ribosomal 28s and 18s bands were determined by agarose-gel electrophoresis.
|
| Label |
Biotin.
|
| Label protocol |
Synthesis of biotin labeled cRNA was performed with BioArray HighYield RNA TranscriptLabeling Kit (ENZO) following standard procedures.
|
| |
|
| Hybridization protocol |
Hybridization and washing was performed following standard procedures for Affymetrix HG-U133A arrays (www.affymetrix.com) on a GeneChip Fluidic station 400, using the EukGe-WS2v4 protocol.
|
| Scan protocol |
Scanning was performed twice with filter set at 570 nm on an Agilent gene array scanner (Affymetrix).
|
| Description |
Sufficient amount of high quality RNA was isolated and infDC transcriptome was assessed using the HG-U133A Affymetrix microarray.
|
| Data processing |
MAS5.0 (Affymetrix) was used for raw data measurement and quality control. dChip 2006 was used for data normalization, quality control and data analysis.
|
| |
|
| Submission date |
Oct 19, 2008 |
| Last update date |
Sep 01, 2016 |
| Contact name |
Joachim Schultze |
| E-mail(s) |
j.schultze@uni-bonn.de
|
| Organization name |
LIMES (Life and Medical Sciences Center Genomics and Immunoregulation)
|
| Department |
Genomics and Immunoregulation
|
| Street address |
Carl-Troll-Strasse 31
|
| City |
Bonn |
| State/province |
NRW |
| ZIP/Postal code |
53115 |
| Country |
Germany |
| |
|
| Platform ID |
GPL96 |
| Series (1) |
| GSE9946 |
Comparison of stimulatory and inhibitory dendritic cell subsets reveals new role of DC in granulomatous infection |
|
| Relations |
| Reanalyzed by |
GSE86363 |
