|
| Status |
Public on May 15, 2018 |
| Title |
CD4 T cells stimulated with Wt APC |
| Sample type |
RNA |
| |
|
| Source name |
spleen
|
| Organism |
Mus musculus |
| Characteristics |
cell type: CD4 T cells
treatment: stimulated with Wt APC
background strain: C57/Bl6
age: 6 weeks-old
|
| Treatment protocol |
Wild type (Wt) B6 and OX40L-Tg APCs (CD3‒ splenocytes) were prepared by depleting T cells from the splenocytes with PE–anti-CD3 (clone 2C11) and anti-PE microbeads (Miltenyi Biotec), followed by a brief treatment with mitomycin C (50μg/ml; Sigma-Aldrich). Naive CD4+ T cells were activated in vitro with soluble anti-CD3 mAb and mitomycin C-treated Wt APCs or OX40L-Tg APCs in 96-well round bottom tissue-culture plates. Forty-eight hours later, living CD4+ T cells were sorted from cultures by flow cytometry.
|
| Growth protocol |
Naive CD4+ T cells were FACS sorted from 6 weeks old Foxp3-eGFP reporter mice with FACSaria (BD Biosciences).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was prepared from mouse adipose tissue using TRIzol reagent and whereafter purified total RNA was isolated using Qiagen RNEasy columns. RNA integrity was checked on chip analysis (Agilent 2100 bioanalyzer, Agilent Technologies, Amsterdam, the Netherlands) according to the manufacturer's instructions. RNA was judged as suitable for array hybridization only if samples exhibited intact bands corresponding to the 18S and 28S ribosomal RNA subunits, and displayed no chromosomal peaks or RNA degradation products (RNA Integrity Number > 8.0).
|
| Label |
biotin
|
| Label protocol |
All target labeling reagents were purchased from Affymetrix (Santa Clara, CA). Double-stranded cDNAs were synthesized from 1ug total RNA through reverse transcription with an oligo-dT primer containing the T7 RNA polymerase promoter and double strand conversion using the cDNA Synthesis System. Biotin-labeled cRNA was generated from the cDNA and used to probe Affymetrix Mouse Genome HT_MG-430 PM arrays.
|
| |
|
| Hybridization protocol |
The GeneChip HT MG-430 PM 24 Array Plate consists of 24 single MG-430 PM arrays arranged into standard 96 well plate format. Array hybridization, washing and scanning were performed according to the manufacturer’s recommendations on a GeneTitan Instrument.
|
| Scan protocol |
Arrays were scanned on an Affymetrix GeneTitan instrument.
|
| Data processing |
Expression estimates were calculated using RMA in Bioconductor.
|
| |
|
| Submission date |
Oct 05, 2017 |
| Last update date |
May 17, 2018 |
| Contact name |
Xiaolong Zhang |
| E-mail(s) |
smdragon@mail.ustc.edu.cn
|
| Organization name |
Houston Methodist Research Institute
|
| Department |
Immunobiology & Transplant Research Center
|
| Street address |
6670 Bertner Avenue, R7-211
|
| City |
HOUSTON |
| State/province |
Texas |
| ZIP/Postal code |
77030 |
| Country |
USA |
| |
|
| Platform ID |
GPL11180 |
| Series (1) |
| GSE104611 |
Expression data from C57/Bl6 Wt mice CD4+ T cells after activation with anti-CD3 plus Wt APC or OX40L-Tg APC |
|
