|
| Status |
Public on May 02, 2007 |
| Title |
AR2-MB-24-2-611 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
midbrain (collagen)
|
| Organism |
Mus musculus |
| Characteristics |
Mid Brain from male DBA/1 mice 31 d after i.d. injection of Type II collagen in Complete Freunds adjuvant
|
| Treatment protocol |
Eight week old DBA/1 male mice were injected i.d. with 100 ul type II Collagen in 100 ul of Complete Freunds ajuvant at day 24 were injected with LPS and sacrifice by decapitation at day 31 h
|
| Growth protocol |
Mice were housed in HEPA filtered air racks (Tecniplast, Italy) with food and water ad libitum
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using TRIzol Reagent (Invitrogen)
|
| Label |
Cy3
|
| Label protocol |
RNA was reversed transcribed using Superscript II RT (Invitrogen) with oligo dT primers and random primers, both in the presence of aa-dUTP (Sigma). The cDNA was labeled with Cy3 or Cy5 dyes (Amersham), resuspended in DMSO, and incubated for 1 hr at room temperature in the dark. The probes were purified using the SNAP Gel Purification Kit (Invitrogen) following manufacturer instructions with the following modification: at the initial step, 500 ul of loading buffer (2.25 M guanidinium HCl in 70% isopropanol) were added to the sample, placed in a SNAP column and incubated for 4 min before the first centrifugation
|
| |
|
| Channel 2 |
| Source name |
midbrain (control)
|
| Organism |
Mus musculus |
| Characteristics |
Mid Brain from male DBA/1 mice 31 d after i.d. injection of Complete Freunds adjuvant
|
| Treatment protocol |
Eight week old DBA/1 male mice were injected i.d. with 200 ul of Complete Freunds ajuvant at day 24 were injected with LPS and sacrifice by decapitation at day 31 h
|
| Growth protocol |
Mice were housed in HEPA filtered air racks (Tecniplast, Italy) with food and water ad libitum
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using TRIzol Reagent (Invitrogen)
|
| Label |
Cy5
|
| Label protocol |
RNA was reversed transcribed using Superscript II RT (Invitrogen) with oligo dT primers and random primers, both in the presence of aa-dUTP (Sigma). The cDNA was labeled with Cy3 or Cy5 dyes (Amersham), resuspended in DMSO, and incubated for 1 hr at room temperature in the dark. The probes were purified using the SNAP Gel Purification Kit (Invitrogen) following manufacturer instructions with the following modification: at the initial step, 500 ul of loading buffer (2.25 M guanidinium HCl in 70% isopropanol) were added to the sample, placed in a SNAP column and incubated for 4 min before the first centrifugation
|
| |
|
| |
| Hybridization protocol |
Slides were prehybridised at 42°C in 2X SSC, 0.1% SDS, 1% BSA. Labeled probes were mixed with hybridization buffer containing 30% formamide and hybridized overnight at 42°C
|
| Scan protocol |
Images were obtained with a VersArray ChipReader system (BioRad) at 10 um resolution and 16bit depth. Each slide was scanned three times, each time with a 5% increment in the sensitivity of the scanner. The data integration for multipole scanning were calculated by M = log2(cy3 / cy2) and A = 0.5 * log2(cy2 * cy3) values for all the possible combinations between channels among the three scannings. Quality score for each couple of channels was calculated as QS = min(QScy2, QScy3). The resulting 9 sets of A and M values were averaged using the QS as weights to obtain the integrated A and M sets.
|
| Description |
Mid Brain of male DBA/1 mice
|
| Data processing |
Data was log2 transformed and normalizaed with an intensity- and location-dependent loess fit
|
| |
|
| Submission date |
Mar 14, 2007 |
| Last update date |
May 04, 2007 |
| Contact name |
Mariano Cesar Salibe |
| E-mail(s) |
msalibe@leloir.org.ar
|
| Organization name |
Gentron LLC
|
| Street address |
Av. Patricias Argentinas 435
|
| City |
Ciudad de Buenos Aires |
| ZIP/Postal code |
1425 |
| Country |
Argentina |
| |
|
| Platform ID |
GPL4987 |
| Series (2) |
| GSE7284 |
profiling of midbrain from arthritic DBA/1 mice vs. control DBA/1 mice |
| GSE7565 |
Profiling brain and liver tissues from mice |
|
