| 1. General features |
| ID #: |
| 768 |
| Name: |
| DIGE analysis for 24hr samples |
| Last update: |
| 2/9/2011 11:55:54 AM |
| |
| 1.1 Responsible person or role |
| Name: |
| Jenna |
| Last name: |
| Kenyani |
| Email: |
| j.kenyani@liverpool.ac.uk |
| Address: |
| School of Veterinary science |
| Locality: |
| Brownlow Hill, Liverpool |
| Country: |
| Uk |
| Institution: |
| University of Liverpool |
| Position: |
| researcher |
| |
| 1.2 Electrophoresis type: |
| difference gel electrophoresis |
| |
| 1.3 Electrophoresis context: |
| MIAPE GE reference: 1082 |
| Link to the report: http://estrellapolar.cnb.csic.es/proteored/MIAPE/MIAPE_GE.asp?pmCodigoAcceso=415db6c1&pmIDUsuario=2378&pmId=1082 |
| |
| 1.4 Image
(1/13) |
| ID: |
| 2037 |
| Name: |
| Gel1-Cy2_internalstandard |
| Type |
| DIGE scans |
| Format: |
| Tagged Image File Format |
| Dimensions: |
| X: 2357 pixels |
| Y: 1807 pixels |
| Resolution: |
| 252 dots per inch |
| Bit-depth: |
| 16 |
| Standard image orientation: |
| Normal orientation |
| Stored in: |
| http://estrellapolar.cnb.csic.es/proteored/MIAPE/Ficheros/462/61713_b_crop_STANDARD_CY2.gel |
| |
| 1.4 Image
(2/13) |
| ID: |
| 2038 |
| Name: |
| Gel1-Cy3-Treated |
| Type |
| DIGE scans |
| Format: |
| Tagged Image File Format |
| Dimensions: |
| X: 2357 pixels |
| Y: 1807 pixels |
| Resolution: |
| 252 dots per inch |
| Bit-depth: |
| 16 |
| Standard image orientation: |
| Normal orientation |
| Stored in: |
| http://estrellapolar.cnb.csic.es/proteored/MIAPE/Ficheros/462/61713_b_crop_CY3.gel |
| |
| 1.4 Image
(3/13) |
| ID: |
| 2039 |
| Name: |
| Gel1-Cy5-Control |
| Type |
| DIGE scans |
| Format: |
| Tagged Image File Format |
| Dimensions: |
| X: 2393 pixels |
| Y: 1782 pixels |
| Resolution: |
| 252 dots per inch |
| Bit-depth: |
| 16 |
| Standard image orientation: |
| Normal orientation |
| Stored in: |
| http://estrellapolar.cnb.csic.es/proteored/MIAPE/Ficheros/462/61713_b_crop_CY5.gel |
| |
| 1.4 Image
(4/13) |
| ID: |
| 2040 |
| Name: |
| Gel2-Cy2_internalstandard |
| Type |
| DIGE scans |
| Format: |
| Tagged Image File Format |
| Dimensions: |
| X: 2429 pixels |
| Y: 1825 pixels |
| Resolution: |
| 252 dots per inch |
| Bit-depth: |
| 16 |
| Standard image orientation: |
| Normal orientation |
| Stored in: |
| http://estrellapolar.cnb.csic.es/proteored/MIAPE/Ficheros/462/61714_crop_STANDARD_CY2.gel |
| |
| 1.4 Image
(5/13) |
| ID: |
| 2041 |
| Name: |
| Gel2-Cy3-treated |
| Type |
| DIGE scans |
| Format: |
| Tagged Image File Format |
| Dimensions: |
| X: 2429 pixels |
| Y: 1825 pixels |
| Resolution: |
| 252 dots per inch |
| Bit-depth: |
| 16 |
| Standard image orientation: |
| Normal orientation |
| Stored in: |
| http://estrellapolar.cnb.csic.es/proteored/MIAPE/Ficheros/462/61714_crop_CY3.gel |
| |
| 1.4 Image
(6/13) |
| ID: |
| 2042 |
| Name: |
| Gel2-Cy5-Control |
| Type |
| DIGE scans |
| Format: |
| Tagged Image File Format |
| Dimensions: |
| X: 2429 pixels |
| Y: 1825 pixels |
| Resolution: |
| 252 dots per inch |
| Bit-depth: |
| 16 |
| Standard image orientation: |
| Normal orientation |
| Stored in: |
| http://estrellapolar.cnb.csic.es/proteored/MIAPE/Ficheros/462/61714_crop_CY5.gel |
| |
| 1.4 Image
(7/13) |
| ID: |
| 2043 |
| Name: |
| Gel3-Cy2_internalstandard |
| Type |
| DIGE scans |
| Format: |
| Tagged Image File Format |
| Dimensions: |
| X: 2471 pixels |
| Y: 1800 pixels |
| Resolution: |
| 252 dots per inch |
| Bit-depth: |
| 16 |
| Standard image orientation: |
| Normal orientation |
| Stored in: |
| http://estrellapolar.cnb.csic.es/proteored/MIAPE/Ficheros/462/61715_STANDARD_CY2(1).gel |
| |
| 1.4 Image
(8/13) |
| ID: |
| 2044 |
| Name: |
| Gel3-Cy5-Treated |
| Type |
| DIGE scans |
| Format: |
| Tagged Image File Format |
| Dimensions: |
| X: 2471 pixels |
| Y: 1800 pixels |
| Resolution: |
| 252 dots per inch |
| Bit-depth: |
| 16 |
| Standard image orientation: |
| Normal orientation |
| Stored in: |
| http://estrellapolar.cnb.csic.es/proteored/MIAPE/Ficheros/462/61715_CY5(1).gel |
| |
| 1.4 Image
(9/13) |
| ID: |
| 2045 |
| Name: |
| Gel3-Cy3-Control |
| Type |
| DIGE scans |
| Format: |
| Tagged Image File Format |
| Dimensions: |
| X: 2471 pixels |
| Y: 1800 pixels |
| Resolution: |
| 252 dots per inch |
| Bit-depth: |
| 16 |
| Standard image orientation: |
| Normal orientation |
| Stored in: |
| http://estrellapolar.cnb.csic.es/proteored/MIAPE/Ficheros/462/61715_CY3(1).gel |
| |
| 1.4 Image
(10/13) |
| ID: |
| 2046 |
| Name: |
| Gel4-Cy2_internalstandard |
| Type |
| DIGE scans |
| Format: |
| Tagged Image File Format |
| Dimensions: |
| X: 2450 pixels |
| Y: 1850 pixels |
| Resolution: |
| 252 dots per inch |
| Bit-depth: |
| 16 |
| Standard image orientation: |
| Normal orientation |
| Stored in: |
| http://estrellapolar.cnb.csic.es/proteored/MIAPE/Ficheros/462/61715_STANDARD_CY2(2).gel |
| |
| 1.4 Image
(11/13) |
| ID: |
| 2047 |
| Name: |
| Gel4-Cy3-Control |
| Type |
| DIGE scans |
| Format: |
| Tagged Image File Format |
| Dimensions: |
| X: 2450 pixels |
| Y: 1850 pixels |
| Resolution: |
| 252 dots per inch |
| Bit-depth: |
| 16 |
| Standard image orientation: |
| Normal orientation |
| Stored in: |
| http://estrellapolar.cnb.csic.es/proteored/MIAPE/Ficheros/462/61716_crop_CY3.gel |
| |
| 1.4 Image
(12/13) |
| ID: |
| 2048 |
| Name: |
| Gel4-Cy5-Treated |
| Type |
| DIGE scans |
| Format: |
| Tagged Image File Format |
| Dimensions: |
| X: 2450 pixels |
| Y: 1850 pixels |
| Resolution: |
| 252 dots per inch |
| Bit-depth: |
| 16 |
| Standard image orientation: |
| Normal orientation |
| Stored in: |
| http://estrellapolar.cnb.csic.es/proteored/MIAPE/Ficheros/462/61716_crop_CY5.gel |
| |
| 1.4 Image
(13/13) |
| ID: |
| 2049 |
| Name: |
| PickGel_Sypro |
| Type |
| DIGE scans |
| Format: |
| Tagged Image File Format |
| Dimensions: |
| X: 2520 pixels |
| Y: 1800 pixels |
| Resolution: |
| 252 dots per inch |
| Bit-depth: |
| 16 |
| Standard image orientation: |
| Normal orientation |
| Stored in: |
| http://estrellapolar.cnb.csic.es/proteored/MIAPE/Ficheros/462/PG2_crop.jpg |
| |
| 1.5 Image analysis software |
| DeCyder - Version: 6.5 |
| Vendor: |
| GE Healthcare Life Sciences |
| Description: |
DIA module (Differencial in Gel Analysis)
BVA module (Biological Variation Analysis) |
| |
| 2. Gel analysis design |
| ID: 561 |
| Name: |
| Design card |
| 2.1 Type: |
| Directed, 2 groups |
| 2.2 Replicates: |
| 4 Biological replicates |
| 2.3 Groups: |
Control1 Cy3 - Treatment1 Cy5 - IntStand1 Cy2
Control2 Cy3 - Treatment2 Cy5 - IntStand2 Cy2
Control3 Cy5 - Treatment3 Cy3 - IntStand3 Cy2
Control4 Cy5 - Treatment4 Cy3 - IntStand4 Cy2
Grouped according to Control or Treatment |
| 2.4 Standards: |
| internal standards - an aditional sample run in the same physical gel to normalize results, from an equal pool of control and treated |
| |
| 3. Image preparation |
| |
| 3.2 Image preparation step |
| Name: |
| Cropping step |
| Type: |
| Cropping- To remove strip at top of image so not to interfer with spot detection |
| Step number: |
| 1st |
|
|
| |
| 4. Image processing |
| Name: |
| Loading images in to DeCyder |
| 4.1 Input image reference (from section 1): |
| ID:1533- Name:Gel1-Cy2_internalstandard |
| ID:1534- Name:Gel1-Cy3-Control |
| ID:1535- Name:Gel1-Cy5-Treated |
| ID:1536- Name:Gel2-Cy2_internalstandard |
| ID:1537- Name:Gel2-Cy3-Control |
| ID:1538- Name:Gel2-Cy5-Treated |
| ID:1539- Name:Gel3-Cy2_internalstandard |
| ID:1540- Name:Gel3-Cy5-Control |
| ID:1541- Name:Gel3-Cy3-treated |
| ID:1542- Name:Gel4-Cy2_internalstandard |
| ID:1543- Name:Gel4-Cy3-treated |
| ID:1544- Name:Gel4-Cy5-Control |
| ID:1545- Name:PickGel_Sypro |
| |
| 4.2 Image processing software |
| DeCyder |
| Vendor: |
| GE Healthcare |
| Description: |
| Image loader module |
| |
| 4.3 Processing step |
| Name: |
| Loading of images |
| Step number: |
| 2nd |
| |
| 5. Data extraction process |
| Name: |
| Data extraction |
| |
| 5.1 Feature detection |
| Algorithm name/version: |
| Spot detection |
| Step number: |
| 3rd |
| Feature editing: |
| Automaticaly in DIA module |
| |
| 5.2 Matching |
| Algorithm name/version: |
| Spot matching |
| Feature reference image used (from section 1): |
| ID:1539- Name:Gel3-Cy2_internalstandard |
| Landmarks: |
| Manually assigned |
| Match editing: |
| Automaticaly in BVA molule and reviewed manualy |
| |
| 5.3 Feature quantitation |
| Algorithm name/version: |
| Quantitation in DeCyder |
| Type: |
| ratios of normalized volumes |
| |
| 5.3.1 Quantitation
|
| DeCyder™ DIA algorithm |
| Parameters: |
| co-detection, background subtraction, normalization, and quantitation of spots in images |
| |
| 5.3.3 Normalization
|
| DeCyder™ DIA algorithm |
| Parameters: |
| co-detection, background subtraction, normalization, and quantitation of spots in images |
| |
| 6. Data analysis |
| Name: |
| Analysis |
| Analysis intent: |
| Finding the spots with a statistical significant differential expression |
| Type: |
| student’s T test was performed and averaged ratios |
| Parameters: |
| Any protein spots which appeared in nine of the twelve gels and had a +/- fold change of 1.5 or above (p<0.05) was highlighted as a spot of interest. |
| |
| 6.2 Data analysis software |
| DeCyder |
| Vendor: |
| GE Healthcare |
| Description: |
| BVA protein statistics |
