|
| Status |
Public on Aug 25, 2007 |
| Title |
p414 sample_Ovarian tumor |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
Total RNA extracted from frozen biopsies of ovarian tumors
|
| Organism |
Homo sapiens |
| Characteristics |
Gender: Female
Status:alive no evidence of disease
FIGO Stage:1
FIGO Substage:c
Histotype:endometrioid
Grade:2
Relapsed:no
Age(years):49.19
Overall Survival(days):1800
Progression Free Survival(days):1800
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA purification from human biopsies were performed using the SV Total RNA Isolation System -cat:#z3100 from Promega.
|
| Label |
Cy5
|
| Label protocol |
RNA amplification for array analysis was performed using Ambion MessageAmp aRNA Kit, cat 1750. cDNA labeling was performed using Amino Allyl cDNA Labeling Kit - cat 1705 by Ambion.
|
| |
|
| Channel 2 |
| Source name |
Clontech Universal Reference
|
| Organism |
Homo sapiens |
| Characteristics |
Human Atlas high pure control total RNA extracted from a pool of 60 human cell lines (BD Biosciences)
|
| Extracted molecule |
total RNA |
| Extraction protocol |
see the producer web site: http//clontech.com/products/detail.asp?tabno=2&catalog_id=636538&page=all&faq_id=153665
|
| Label |
Cy3
|
| Label protocol |
RNA amplification for array analysis was performed using Ambion MessageAmp aRNA Kit, cat 1750. cDNA labeling was performed using Amino Allyl cDNA Labeling Kit - cat 1705 by Ambion.
|
| |
|
| |
| Hybridization protocol |
Microarray hybridization: Add to each Sample 2.5 ul Cot-1 DNA and 12.50 ul 2X Hybridization Buffer. Heat samples for 2 min at 65 C. Centrifuge. Pipet the cDNA array and 22 ul of sample. Incubate for 16 hr at 65 C. Used Atlas Glass Microarray User Manual PT3453-1 instructions for post-hybridization wash.
|
| Scan protocol |
Arrays were scanned using a laser-scanner Axon 4000-B, (Axon Instruments, Union City, CA), using 5-micron resolution. Within the GenePix program were chosen photo-multiplier parameters that defined the quanta intensity density distribution of the array to give a Cy3/Cy5 ratio close to 1.0 and produced not too much saturated signals for each array. Separate 16-bit images were acquired for Cy3 and Cy5.
|
| Description |
none
|
| Data processing |
Image addressing and segmentation was performed using CSIRO Spot software, and data analysis occurred with R statistical software package. For each found array dataset two distinct classes of log2 ratio have been computed. The first (MA=(FR BGR)/(FG BGG)) was the classic log ratio of the two net fluorescence signal (F-BG) determined by subtraction of the local background (BG) from the spot median intensity (F). The second (MB=(FR/BGR)/(FG/BGG)) instead was the log ratio of the two relative intensities (F/BG) of each channel fluorescence (F) of spot in respect to its own local background (BG). The log2 geometric mean intensities and log ratios of both classes were arbitrarily set to 0 if a spot had not a net fluorescence signal greater than 0 in both channels (i. e. is a unreliable spot). Both kind of M-values were normalised and corrected from possible biases using the LOWESS. In order to evaluate which genes have probably modified their expression level an intensity-dependent Z-score calculation has been performed for each microarray data set and log ratio class (MA and MB).
|
| |
|
| Submission date |
Jul 31, 2007 |
| Last update date |
Aug 14, 2011 |
| Contact name |
Pietro Mariani |
| Organization name |
Mario Negri Institute
|
| Street address |
Via La Masa 19
|
| City |
Milan |
| ZIP/Postal code |
20156 |
| Country |
Italy |
| |
|
| Platform ID |
GPL5689 |
| Series (1) |
| GSE8841 |
ANALYSIS OF GENE EXPRESSION IN EARLY-STAGE OVARIAN CANCER (1) |
|
