|
| Status |
Public on Dec 20, 2012 |
| Title |
PBMC_C4-8w_rep1 |
| Sample type |
RNA |
| |
|
| Source name |
Control_PBMC_8 wk
|
| Organism |
Homo sapiens |
| Characteristics |
status: control
gender: male
age: 37
treatment: No treatment
sample collection: 8 weeks later after clinical remission
|
| Extracted molecule |
total RNA |
| Extraction protocol |
8-10ml venous peripheral blood from fasting patients and healthy controls were collected during 7am and 9am. PBMCs were separeted by Ficoll gradient centrifugation.Total RNA was isolated using the mirVanai Kit (Ambion) according to manufacturer's recommendation. RNA was quantified using a NanoDrop-1000 spectrophotometer and quality was monitored with the Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA).
|
| Label |
Cy3
|
| Label protocol |
Cyanine-3 (Cy3) labeled cRNA was prepared from 200ng RNA using the One-Color Low RNA Input Linear Amplification PLUS kit (Agilent) according to the manufacturer's instructions, followed by RNAeasy column purification (QIAGEN, Valencia, CA). Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.
|
| |
|
| Hybridization protocol |
600 ng of Cy3-labelled cRNA (specific activity > 6 pmol Cy3/ug cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 25ul containing 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturers instructions. On completion of the fragmentation reaction, 250 ml of 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to Agilent Whole Human Genome Oligo Microarrays (G4851A) for 17 hours at 65°C in a rotating Agilent hybridization oven. After hybridization, microarrays were washed 1 minute at room temperature with GE Wash Buffer 1 (Agilent) and 1 minute with 37°C GE Wash buffer 2 (Agilent), then dried immediately by brief centrifugation.
|
| Scan protocol |
Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner (G2505C) using one color scan setting for 8x60k array slides.
|
| Description |
Gene expression of control PBMCs
|
| Data processing |
The scanned images were analyzed with Feature Extraction Software 10.5.1.1 (Agilent) using default parameters (protocol GE1_107_Sep09 and Grid: 028004_D_F_20110325) to obtain background subtracted and spatially detrended Processed Signal intensities. All data were normalized by quantile normalization using limma R/bioconductor package (v.2.16.4).
|
| |
|
| Submission date |
May 24, 2012 |
| Last update date |
Dec 20, 2012 |
| Contact name |
El Chérif Ibrahim |
| E-mail(s) |
el-cherif.ibrahim@univ-amu.fr
|
| Phone |
+33 (0)4 91 69 89 56
|
| Fax |
+33 (0)4 91 69 89 20
|
| Organization name |
Aix Marseille Université-CNRS
|
| Department |
CRN2M-UMR7286
|
| Street address |
Bd Pierre Dramard
|
| City |
MARSEILLE |
| ZIP/Postal code |
13015 |
| Country |
France |
| |
|
| Platform ID |
GPL13607 |
| Series (1) |
| GSE38206 |
Transcription profiling of major depression peripheral blood mononuclear cells (PBMCs) at clinical remission compared to severe acute state |
|
