|
| Status |
Public on Dec 01, 2011 |
| Title |
Ctrl4 |
| Sample type |
RNA |
| |
|
| Source name |
microvesicle RNA, healthy control
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: blood
cell type: serum microvesicles
disease state: healthy
|
| Treatment protocol |
Blood samples were collected into a BD Vacutainer SST (#367985). The blood was left to clot for 30 min and serum was isolated according to manufacturer’s recommendations within two hours of collection. Serum was filtered by slowly passing it through a 0.8 µm syringe filter, aliquoted into 1.8 mL cryotubes and kept at -80°C until used.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
One mL of serum was transferred to an ultracentrifuge tube, diluted 1:1 with cold PBS and centrifuged at 110,000g for 70 min at 4°C. The supernatant was carefully aspirated off without disturbing the microvesicle pellet. The pellet was treated for 15 min with 50 μL DNase (Ambion), 700 μL miRNeasy lysis buffer was added to the tube and the RNA was isolated following the manufacturer’s recommendations. After elution of the RNA from the column in water, the RNA was precipitated by adding 100% EtOH and incubation at -20°C, centrifugation and removal of the supernatant. The pellet was left to dry and dissolved in 14 μL of nuclease-free water and stored at -80°C until needed.
|
| Label |
Cy3
|
| Label protocol |
Linear amplification was carried out by Miltenyi according to manufacturer’s recommendations. Briefly, exoRNA was linearly amplified and fluorescently labeled with Cy3 using [XXX Kit Name] (Agilent).
|
| |
|
| Hybridization protocol |
Hybridization to Agilent microarrays was carried out by Miltenyi according to manufacturer’s recommendations. Briefly, 1.4 µg amplified RNA was hybridized to Agilent 4x44K Human Microarrays and the arrays were washed.
|
| Scan protocol |
The arrays were scanned on an Agilent Scanner G2505C. Raw data was generated by image analysis using Feature Extraction version 10.5.1.1 (Agilent).
|
| Data processing |
The data were normalized.
|
| |
|
| Submission date |
Sep 10, 2010 |
| Last update date |
Dec 01, 2011 |
| Contact name |
Mikkel Noerholm |
| Organization name |
MGH
|
| Department |
Neurology
|
| Lab |
Breakefield
|
| Street address |
149 13th Street
|
| City |
Charlestown |
| State/province |
MA |
| ZIP/Postal code |
02129 |
| Country |
USA |
| |
|
| Platform ID |
GPL8755 |
| Series (1) |
| GSE24084 |
RNA expression patterns in serum microvesicles from patients with glioblastoma multiforme and controls |
|
