|
| Status |
Public on Mar 10, 2012 |
| Title |
T2M cells_replicate 1 |
| Sample type |
RNA |
| |
|
| Source name |
T2M
|
| Organism |
Mus musculus |
| Characteristics |
cell surface markers: IL-4/GFP+ IL-5rα- F4/80+ MHCIIlo CD11b+ Gr-1mid
background strain: BALB/c
genotype/variation: 4get
tissue type: lung
cell type: T2M
|
| Treatment protocol |
Lung tissue was processed via enzymatic digestion with 1 mg/ml collagenase A (Roche) in RPMI 1640 (Invitrogen) with 5% FCS (Atlas Biologicals) and 2 U DNase (Sigma-Aldrich) at 37°C for 1 hour, and cells dispersed by passing digest through an 18-gauge cannula. Following RBC lysis,
MACS columns (Miltenyl) were used to enrich for CD11b+ myleoid populations. Cells were pooled from 4 donor mice and stained for flow cytometry using standard techniques. Cells surface markers used to discriminate between inflammatory subsets are listed in the sample description column.
|
| Growth protocol |
6–8 week old female 4get mice received daily intra-tracheal injections (0.5 μg recombinant IL-25 (R&D) in 50 μL PBS) for 4 days to induce antigen-independent type 2 inflammation.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
400,000 cells were collected by FACS for each sample and lysed directly upon sorting in RLT lysis buffer. Total RNA was isolated with Qiagen kits according to manufacturer's instructions.
|
| Label |
biotin
|
| Label protocol |
Biotinylated cDNA were prepared according to the Nugen Ovation V2 kit protocol from 50 ng total RNA (ovation pico WTA system V2. PN3302)
|
| |
|
| Hybridization protocol |
Following Labeling, 4 ug of cDNA were hybridized for 16 hr at 45C on GeneChip Mouse 430 2.0 Array. GeneChips were washed and stained in the Affymetrix Fluidics Station 450.
|
| Scan protocol |
GeneChips were scanned using the Affymetrix 3000 7G GeneChip Scanner with Autoloader
|
| Description |
Lung, IL-25 treated
|
| Data processing |
Data was analyzed using the limma and affy Bioconductor packages implemented in the R-statistical environment. RMA was used to normalize the data and fit log2 transformed expression values.
|
| |
|
| Submission date |
Mar 09, 2012 |
| Last update date |
Mar 10, 2012 |
| Contact name |
Bryan C Petersen |
| E-mail(s) |
[email protected]
|
| Organization name |
University of Michigan
|
| Department |
Pathology
|
| Street address |
109 Zina Pitcher, 4620 BSRB
|
| City |
Ann Arbor |
| State/province |
MI |
| ZIP/Postal code |
48103 |
| Country |
USA |
| |
|
| Platform ID |
GPL1261 |
| Series (1) |
| GSE36392 |
Expression data from pulmonary Type 2 Myeloid (T2M) cells, Eosinophils, Neutrophils, and Macrophages, from IL-25 treated 4get mice |
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