|
| Status |
Public on Sep 06, 2008 |
| Title |
Primary ovine microglia PrPSc bio repC tech rep3 (Inoc12C.3) |
| Sample type |
RNA |
| |
|
| Source name |
primary ovine microglial cells inoculated with PrPSc
|
| Organism |
Ovis aries |
| Characteristics |
Primary ovine microglia
|
| Treatment protocol |
Primary microglia were passed into 6-well plates and grown to approximately 60% confluency. Microglia were rinsed once with PBS and then overlaid with 200 μl of a 1/20 dilution of either the Rov9Sc lysate (Inoc A, B, and C) or the Rov9C lysate (Mock A, B, and C) in OPTI-MEM. All replicates (A, B, and C) of both treatment groups (Inoc and Mock) were incubated for six hours and then 200 μl of OMEM were added to each well. Following an additional two days of incubation, 0.5 ml of OMEM was added to each well, and microglia were incubated for four days at which time they were expanded into 25-cm2 tissue culture flasks. Microglia were fed every three to four days with OMEM as necessary and serially passed 1/5 after reaching confluence.
|
| Growth protocol |
Microglial cells were cultured using standard techniques, in OPTI-MEM I Reduced Serum Medium (Invitrogen, Carlsbad, CA) supplemented with 10% heat-inactivated FBS, 2 mM L-glutamine, 10 IU/ml of penicillin, and 10 mg/ml of streptomycin.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
QIAshredder spin columns (Qiagen, Valencia, CA) followed by Qiagen RNeasy mini spin columns
|
| Label |
biotin
|
| Label protocol |
10ug total RNA labelled according to Affymetrix one cycle labelling protocol
|
| |
|
| Hybridization protocol |
Affymetrix hybridization protocols; Affymetrix GeneChip Fluidics Station 450
|
| Scan protocol |
Affymetrix GeneChip Scanner 300 7G
|
| Description |
PrPSc infected primary ovine microglial cells
|
| Data processing |
All analyses of microarray data were conducted using Bioconductor for R (www.R-project.org). Data were normalized using the Micro Array Suite 5.0 (MAS5) algorithm. The ComBat algorithm (http://statistics.byu.edu/johnson/ComBat/) was used to correct for batch effect.
|
| |
|
| Submission date |
Sep 05, 2008 |
| Last update date |
Sep 05, 2008 |
| Contact name |
James Stanton |
| Organization name |
Washington State University
|
| Street address |
PO Box 647034
|
| City |
Pullman |
| State/province |
WA |
| ZIP/Postal code |
99164-7034 |
| Country |
USA |
| |
|
| Platform ID |
GPL2112 |
| Series (1) |
| GSE12688 |
Expression profiling of prion accumulating ovine microglia |
|
