|
| Status |
Public on Mar 01, 2011 |
| Title |
PBMC_normal_validation_3 |
| Sample type |
RNA |
| |
|
| Source name |
PBMCs from patients with normal mammogram
|
| Organism |
Homo sapiens |
| Characteristics |
phenotype: Normal
tissue: peripheral blood mononuclear cells
data set: validation
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA from peripheral blood mononuclear cells was isolated using the Ambion Ribopure™-Blood kit (Ambion, Austin, TX).
|
| Label |
biotin
|
| Label protocol |
Biotinylated cRNA were prepared according to the standard Affymetrix IVT 1-cycle protocol from 2.5 ug total RNA.
|
| |
|
| Hybridization protocol |
Following fragmentation, 15ug of labeled cRNA were hybridized on HU133 Plus 2.0 GeneChips for 16 hours at 45°C. GeneChips were washed and stained using the Biomex FX liquid handler.
|
| Scan protocol |
GeneChips were scanned using the Axon Genepix Scanner (Genepix 4000B).
|
| Description |
Blood collection: Blood samples (8mL) collected by venipuncture into BD Vacutainer CPT Cell Preparation tubes. Samples processed within 2 hours of collection.
PBMC isolation: CPT tubes were centrifuged for 30 minutes at 2,500 rpms in a centrifuge with a swinging bucket rotor. The plasma layer was removed, and then the remaining buffy coat was poured into a 15 mL conical tube. Next, 5 mL of chilled phosphate buffered saline (PBS) with 2% fetal bovine serum (FBS) was added to the CPT tube, which was capped and inverted to mix. The contents were poured into the same 15 mL conical tube, which was then centrifuged for 10 minutes at 1200 rpms at room temperature. The supernatant was then aspirated and the pellet was resuspended in 1 mL of freezing media (RPMI with 20% DMSO and 20% FBS). The cells were then transferred to a cryovial (Nunc, Roskilde, Denmark) and placed in a freezer box at -80°C. After 24 hours samples were transferred to liquid nitrogen storage.
|
| Data processing |
Data processing was performed in a pilot revision control system entitled Quality-Auditable Data Repository and Analysis (QUADRA). For fully versioned and annotated source code and data objects, please see http://quadra.genome.duke.edu/mouse-2-human-mammary-tumor-pbmc-project.
Data were RMA normalized according to the source code found at http://quadra.genome.duke.edu/mouse-2-human-mammary-tumor-pbmc-project/human-dataprep.
|
| |
|
| Submission date |
Feb 28, 2011 |
| Last update date |
Nov 14, 2018 |
| Contact name |
Erich Huang |
| E-mail(s) |
[email protected]
|
| Phone |
919-684-6849
|
| Fax |
919-681-8973
|
| Organization name |
Duke University
|
| Department |
Institute for Genome Sciences and Policy
|
| Street address |
101 Science Drive
|
| City |
Durham |
| State/province |
NC |
| ZIP/Postal code |
27708 |
| Country |
USA |
| |
|
| Platform ID |
GPL570 |
| Series (2) |
| GSE27562 |
Expression data from human PBMCs from breast cancer patients and controls |
| GSE27567 |
Integrating Factor Analysis and a Transgenic Mouse Model to Reveal a Peripheral Blood Predictor of Breast Tumors |
|
| Relations |
| Reanalyzed by |
GSE122511 |
