|
| Status |
Public on Aug 01, 2015 |
| Title |
HV-4 |
| Sample type |
RNA |
| |
|
| Source name |
Purified B lymphocytes
|
| Organism |
Homo sapiens |
| Characteristics |
disease: HV
cell type: B cell
|
| Treatment protocol |
Venous blood samples were collected in EDTA vacutainers and processed for analysis within 4 hours. Peripheral Blood Mononuclear Cells (PBMC) were separated on a Ficoll layer (Eurobio, Les Ulis, France) and frozen in 10% DMSO diluted human plasma (Sigma-Aldrich, St Louis, MO). Then total B cells were purified from PBMC by negative selection on magnetic columns according to the manufacturer’s instructions (B cell isolation Kit II, Miltenyi Biotech, Gladbach, Germany).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA were extracted and purified from human B using the RNeasy microkit following the manufacturer’s instructions (Qiagen). RNA was quantified using a NanoDrop ND-1000 spectrophotometer (NanoDrop Technologies, Wilmington, DE), and quality was monitored with the Agilent 2100 Bioanalyzer (Agilent Technologies, Palo Alto, CA).
|
| Label |
cy5
|
| Label protocol |
The cRNA labeling and hybridizations were performed according to Protocol from Agilent Technologies Inc (Santa Clara, CA), using the Agilent Quick Amp Labeling Kit (Agilent Technologies, Palo Alto, CA) following the two-colour manufacturer’s protocol. Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.
|
| |
|
| Hybridization protocol |
cRNA samples were hybridized to a Agilent Technologie Human Gene Expression 8x60K Microarray during 17 hours at 65°C in a rotating hybridization oven (Agilent Technologies). The hybridization slides were washed, stabilized, dried, and immediately scanned.
|
| Scan protocol |
Scanned with an Agilent Technologies Microarray Scanner using default parameters (protocol GE2_107_Sep09 and Grid: 028004_D_F_20110819).
|
| Description |
Raw data file: US82400123_252800413491_S01_GE2_107_Sep09_2_3.txt
Sample name: HV4_13491_2_3
|
| Data processing |
Data extraction of median feature intensity without background subtraction was peformed with Feature Extraction software v10.7.1.1 (gMedianSignal or rMedianSignal column)(Agilent Technologies). In order to remove systemic signal intensity bias between each array, median feature intensity were normalized with the function lowess (Locally weighted scatterplot smoothing) method, and then spots with half of the samples for each group exhibiting signal less than the mean of all median signals were removed (threshold: 96.95). 30, 371spots were kept out of 58,717.
Each sample represents one channel of a dual-channel array.
|
| |
|
| Submission date |
Aug 06, 2012 |
| Last update date |
Aug 01, 2015 |
| Contact name |
Richard Danger |
| E-mail(s) |
richard.danger@univ-nantes.fr
|
| Phone |
+33 2 40 08 75 65
|
| Organization name |
CHU Nantes - INSERM- Nantes Universite
|
| Department |
U1064
|
| Lab |
CR2TI
|
| Street address |
30 Boulevard Jean Monnet
|
| City |
Nantes |
| ZIP/Postal code |
44035 |
| Country |
France |
| |
|
| Platform ID |
GPL13607 |
| Series (1) |
| GSE39899 |
Gene expression profiling in B lymphocytes from transplanted patients |
|
