|
| Status |
Public on May 31, 2010 |
| Title |
WT, biological replicate 2 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
alveolar macrophages
|
| Organism |
Mus musculus |
| Characteristics |
strain: C57BL6N
cell type: alveolar macrophages
stimulation: no
|
| Treatment protocol |
10 micromolar serotonin (Sigma Aldrich), for 4h
|
| Growth protocol |
RPMI medium with dialysed 1%FSC, 37ºC, 5%CO2 incubator
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was extracted using the RNeasy Midi Kit (Qiagen, Hilden, Germany) according to the manufacturer’s instructions.
|
| Label |
Cy3
|
| Label protocol |
Purified total RNA was amplified and Cy-labeled using the dual-color QuickAmp kit (Agilent Technologies, Palo Alto, CA) following the kit instructions.
|
| |
|
| Channel 2 |
| Source name |
alveolar macrophages
|
| Organism |
Mus musculus |
| Characteristics |
stimulation: yes
strain: C57BL6N
cell type: alveolar macrophages
|
| Treatment protocol |
10 micromolar serotonin (Sigma Aldrich), for 4h
|
| Growth protocol |
RPMI medium with dialysed 1%FSC, 37ºC, 5%CO2 incubator
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was extracted using the RNeasy Midi Kit (Qiagen, Hilden, Germany) according to the manufacturer’s instructions.
|
| Label |
Cy5
|
| Label protocol |
Purified total RNA was amplified and Cy-labeled using the dual-color QuickAmp kit (Agilent Technologies, Palo Alto, CA) following the kit instructions.
|
| |
|
| |
| Hybridization protocol |
Hybridization was performed at 65°C over-night on Agilent G4122F mouse whole genome microarrays (Agilent Technologies, Böblingen, Germany) as indicated by the supplier.
|
| Scan protocol |
Slides were scanned using the GenePix 4100A scanner (MDS Analytical Technologies, Ismaning, Germany) and analyzed with GenePix software V5.1 (MDS). PMT gains were manually adjusted to get some saturated pixels while matching the intensity histograms of Cy5 and Cy3 fluorescence.
|
| Description |
Biological replicate 2
|
| Data processing |
The experiment contains two data sets of 4 hybridizations each. One set of data was in obtained from 1x44k arrays (GSM476104-GSM476107) and a second set of hybridizations was done on the newer 4x44k arrays (GSM476108-GSM476111).
For the analysis, the data of all eight arrays were assembled into a data structure corresponding to GPL4134. The missing dark corner features and spike-in-features were taken from the GPL4134 arrays only. All other spots were matched by their probe IDs. This data set was then analysed to produce the matrix file. The correlation of spot intensities between the two sets was checked carfully and is within the acceptable margins of error, demonstrating that the two sets of data are in fact comparable and the spot matching worked. Further, the analysis of the individual sets gives very similar results to the combined analysis (limma uses a variance estimate from all arrays, the results are not expected to be identical). The data in the matrix file is the result of the combined analysis, what is to be preferred over the individual analyses since the variance estimates are slightly more robust when more arrays are involved.
MA data was calculated from the Genepix data and lowess-nomalized using R (http://www.R-project.org).
|
| |
|
| Submission date |
Nov 27, 2009 |
| Last update date |
May 31, 2010 |
| Contact name |
Jochen Wilhelm |
| Organization name |
Uniklinikum Giessen
|
| Department |
ECCPS Microarray Unit
|
| Street address |
Gaffkystrasse 10
|
| City |
Giessen |
| ZIP/Postal code |
35392 |
| Country |
Germany |
| |
|
| Platform ID |
GPL4134 |
| Series (1) |
| GSE19214 |
Serotonin stimulation of alveolar macrophages from wild type and 5-HT2C deficient animals |
|
