|
| Status |
Public on Sep 30, 2012 |
| Title |
Foxp3-GFPxAPCΔ468 mesenteric lymph node CD4+ non-Treg, biological rep 3 |
| Sample type |
RNA |
| |
|
| Source name |
mesenteric lymph node CD4+ non-Treg from polyp ridden mice
|
| Organism |
Mus musculus |
| Characteristics |
tissue: Mesenteric lymph node
genetic background: C57BL/6
genotype: Foxp3-GFPxAPCΔ468
age: 4 months old
gender: male
|
| Treatment protocol |
Mice were sacraficed and organs were harvested. For polyp-ridden APC mice, small intestine polyps were microdisected from health surrounding tissue; both parts were used as a source of mononuclear cells. Mononuclear cells from small intestine tissue were isolated by collengenase IV digestion and precoll gradient centrifugation. Cells from spleen and mesenteric lymph nodes were isolated by gentlely teasing between frosted glass slides. Spleen cells were subjected to red blood cell lysis. Negative selection with T-cell negative selection kit (Dynal beads; Invitrogen) was used to pre-purify cells. Cells were Pc blocked and then stained with CD4 antibody. PI was used to exclude dead cells during FACS sorting. Cells were then double sorted on a MoFlo sorter. A doublet exclusion gate was included. Cells were then gated on PI negative, CD4+ cells. Two populations were then collected directly into Trizol: CD4+Foxpe-GFP- and CD4+Foxp3-GFP+. Cell isolation and sorting were performed according to immgen.org SOP.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Trizol
|
| Label |
biotin
|
| Label protocol |
Affymetrix GeneChip WT Sense Target Labeling and Control Reagents
|
| |
|
| Hybridization protocol |
Affymetrix GeneChip WT cDNA Synthesis and Amplification Kits
|
| Scan protocol |
Affymetrix GeneChip Scanner
|
| Data processing |
The datasets were pre-filtered to keep only those probesets for which a gene symbol could be found in the affymetrix annotation. CEL files were normalized using Affymetrix Power tools on the predefined probeset ID list mentioned above, and using the standard RMA workflow (background adjustment, quantiile normalization, median polish probeset summarization).
|
| |
|
| Submission date |
Sep 29, 2012 |
| Last update date |
Sep 30, 2012 |
| Contact name |
Nichole Blatner |
| E-mail(s) |
[email protected]
|
| Phone |
312-503-1905
|
| Fax |
312-503-0386
|
| Organization name |
Northwestern University
|
| Department |
Robert H. Lurie Comprehensive Cancer Center
|
| Lab |
Khazaie Lab
|
| Street address |
303 East Superior Street, Lurie 3-250
|
| City |
Chicago |
| State/province |
IL |
| ZIP/Postal code |
60611 |
| Country |
USA |
| |
|
| Platform ID |
GPL6246 |
| Series (1) |
| GSE41229 |
Expression data from T-cells isolated from healthy mice or mice with polyposis |
|
