|
| Status |
Public on Sep 10, 2011 |
| Title |
101 |
| Sample type |
RNA |
| |
|
| Source name |
251485013912_4
|
| Organism |
Homo sapiens |
| Characteristics |
tumor stage: III
rec.: 0
tissue type: CC
|
| Treatment protocol |
Primary tumor tissue and adjacent normal tissues were collected at the time of surgery and were snap-frozen in liquid nitrogen within 30 min. Histopathological examination following hematoxilin/ eosin staining of tissue sections ensured that only tumor samples containing at least 60% of non necrotic tumor epithelium were used for further analysis.
|
| Growth protocol |
Cryo-conserved primary tumour tissue were collected immediately after surgical removal snap-frozen in liquid nitrogen.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Qiagen Rneasy extraction of total RNA was performed according to the manufactures instructions.
|
| Label |
Cy3
|
| Label protocol |
Cyanine-3 (Cy3) labeled cRNA was prepared from 0.5 ug 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 |
1.5 ug of Cy3-labelled cRNA (specific activity >11.0 pmol Cy3/ug cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 110µl containing 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturers instructions. On completion of the fragmentation reaction, 100 µl of fragmentation mixture were hybridized to Agilent Whole Human Genome Oligo Microarrays (G4112F) 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 (G2565AA) using one color scan setting for 4x44k array slides (Scan Area 61x21.6 mm, Scan resolution 5um, Dye channel is set to Green and Green PMT is set to XDR Hi 100%, XDR Lo 10%).
|
| Data processing |
Extract tif images using the Feature Extraction software FE9.1 by GE1-v5_91 protocols. TXT data files were improted in GeneSpring GX10.0.2 software (Agilent Technologies, Santa Clara, USA) and pre-processed using the agilent algorithm. Baseline transformation to median of all samples
|
| |
|
| Submission date |
Sep 06, 2011 |
| Last update date |
Sep 10, 2011 |
| Contact name |
Qing Wang |
| E-mail(s) |
wang.qing@mdc-berlin.de
|
| Phone |
+493094063798
|
| Fax |
+493094062846
|
| Organization name |
Charite-Berlin
|
| Department |
Surgical Oncology
|
| Street address |
Robert Rössle Str. 10
|
| City |
Berlin |
| State/province |
Berlin |
| ZIP/Postal code |
13125 |
| Country |
Germany |
| |
|
| Platform ID |
GPL6480 |
| Series (1) |
| GSE31905 |
Gene expression profiling of colorectal carcinoma |
|
