|
| Status |
Public on Jan 01, 2011 |
| Title |
IgE-crosslinked BMMC_rep2 |
| Sample type |
RNA |
| |
|
| Source name |
activated BMMCs
|
| Organism |
Mus musculus |
| Characteristics |
strain: Balb/c
group: IgE-crosslinked BMMC
|
| Treatment protocol |
Bone marrow-derived mouse mast cells (BMMCs) were cultured for 4 weeks in the presence of SCF and IL-3. On the 6th day, CD117+ cells (putative committed immature progenitors) were magnetically separated. In order to differentiate the 4 weeks old BMMCs toward mucosal-type mast cells the medium was supplemented with IL-9+TGF-beta for 5 additional days. BMMCs were activated by IgE-crosslinking.
|
| Growth protocol |
Culture of bone marrow cells in complete RPMI 1640 supplemented with IL3 (5ng/ml) and SCF (20ng/ml) for 4 weeks.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNAs from cell samples was extracted by miRNeasy Mini Kit, the quantity and quality of total RNA was assessed by NanoDrop ND-1000 spectrophotometer and Agilent 2100 Bioanalyzer, respectively. Only those samples were used for microarray experiments that gave >8.0 for RNA integrity number, showed a clear gel image and no DNA contamination was observed on the histogram.
|
| Label |
Cy3
|
| Label protocol |
Standard Agilent labeling protocol with 100 ng quality-checked total RNA.
|
| |
|
| Hybridization protocol |
Standard Agilent labeling protocol: the labeled samples were hybridized for 20 hours at 55°C.
|
| Scan protocol |
Standard Agilent labeling protocol: the arrays were scanned with an Agilent DNA Microarray Scanner BA. The scanned images were analyzed with Feature Extraction Software 9.5 (Agilent) using default parameters (protocol miRNA-v1_95_May07 and grid: 019119_D_20080215).
|
| Description |
microRNA profiling of activated mast cells obtained by presensitization of BMMCs with anti-DNP IgE followed by IgE-crosslinking by DNP-antigen challenge for 2 hours
|
| Data processing |
Extracted data were analyzed by Genespring GX10.0. Raw data were normalized to the 75th percentile signal intensity and entities showing present call in all samples of a condition were filtered out. Differentially expressed genes were selected when passing the signal intensity filter (entities where at least 100 percent of samples in any 1 out of 4 conditions have values within cut-off) and showing at least 2-fold statistically significant change (ANOVA and Tukey HSD post-hoc test, with Benjamini-Hochberg Multiple Testing Correction p-value<0,05) between BMMC and any other groups.
|
| |
|
| Submission date |
Sep 23, 2010 |
| Last update date |
Jan 01, 2011 |
| Contact name |
Viktor Molnár |
| E-mail(s) |
molvik.dgci@gmail.com
|
| Organization name |
Semmelweis University
|
| Department |
Department of Genetics, Cell- and Immunobiology
|
| Street address |
Nagyvárad tér 4.
|
| City |
Budapest |
| ZIP/Postal code |
1089 |
| Country |
Hungary |
| |
|
| Platform ID |
GPL8824 |
| Series (1) |
| GSE24321 |
MicroRNA profiling of mouse bone marrow-derived mast cells during mucosal differentiation and IgE-mediated activation |
|
