|
| Status |
Public on Jul 15, 2019 |
| Title |
HIV_specific_CD4_ T cells_EC4 |
| Sample type |
RNA |
| |
|
| Source name |
CD154+ CD69+ HIV-specific CD4 T cells from PBMCs
|
| Organism |
Homo sapiens |
| Characteristics |
disease status: Elite Controller
cell type: PBMCs
|
| Treatment protocol |
A CD40 blocking antibody (Miltenyi Biotec, clone HB14) was added 15 minutes prior to stimulation with a gag peptide pool. Following a 9 hr incubation, HIV-specific CD4 T cells were identified based on the co-upregulation of CD40L and CD69 and were live-sorted directly in RLT lysis buffer (Qiagen).
|
| Growth protocol |
Cryopreserved PBMCs were thawed and rested at 37°C for 3 hours.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was purified using the RNeasy Plus Micro Kit (Qiagen) according to manufacturer's instructions.
|
| Label |
biotin
|
| Label protocol |
Sense-strand cDNA was synthesized from total RNA using a fixed volume of 3µL. Fragmentation and labeling were performed to produce ss-DNA with the Applied Biosystems™ GeneChip® WT Pico Terminal Labeling Kit according to manufacturer’s instructions (Applied Biosystems™).
|
| |
|
| Hybridization protocol |
5 µg DNA target was hybridized on GeneChip® Clariom™ D, human (ThermoFisher Scientific) and incubated at 450C in the Genechip® Hybridization oven 640 (Affymetrix) for 17 hours at 60 rpm. GeneChips were then washed in a GeneChips® Fluidics Station 450 (Affymetrix) using Applied Biosystems™ Hybridization Wash and Stain kit according to the manufacturer’s instructions (Applied Biosystems™)
|
| Scan protocol |
The microarrays were finally scanned on a GeneChip® scanner 3000 (Affymetrix).
|
| Data processing |
The preprocessing of the microarray data was done in R v3.3.3 using the oligo package47 v1.38.0 using rma function.
|
| |
|
| Submission date |
Apr 16, 2019 |
| Last update date |
Jul 16, 2019 |
| Contact name |
Daniel Elias Kaufmann |
| Organization name |
CRCHUM
|
| Street address |
900 Rue St. Denis
|
| City |
Montreal |
| State/province |
Quebec |
| ZIP/Postal code |
H2X 0A9 |
| Country |
Canada |
| |
|
| Platform ID |
GPL23126 |
| Series (2) |
| GSE128297 |
Transcriptional analysis of HIV-specific CD4+ T cells |
| GSE129872 |
Transcriptional analysis of HIV-specific CD4+ T cells in progressive disease and in spontaneously controlled infection. |
|
