|
| Status |
Public on Dec 01, 2023 |
| Title |
CD14+ cDC2s_rep1 |
| Sample type |
RNA |
| |
|
| Source name |
primary CD14+ CD1c+ cDC2s donor 1
|
| Organism |
Homo sapiens |
| Characteristics |
individual: donor 1
tissue: blood
cell type: PBMCs
facs isolated cells: CD14+ cDC2s
disease status: healthy donor
|
| Treatment protocol |
PBMCs were isolated using Lymphoprep (Axis-Shield PoC AS). To obtain separate CD1c+CD14- and CD1c+CD14+ populations, and CD14+ monocytes for RNA analysis, PBMCs treated with FcR blocker and depleted for CD3, CD19, and CD56 using MACS microbeads were stained with anti-CD1c-BV421, anti-CD3/CD56/CD20-FITC, and anti-CD14-APC-H7 for 30 min at 4 °C in sterile polypropylene round-bottom tubes (Falcon). Sorting was performed on a BD FACSAria. After sorting, cells were washed twice with PBS and dry cell pellets were stored at -80 until RNA isolation.
|
| Growth protocol |
cells were not cultured and kept at 4C untill cell pellets were prepared for RNA extraction
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA extraction was performed using the Rnease Plus Micro Kit (Qiagen) according to the manufacturer's protocol. All subsequent protocols were performed by Eurofins.
|
| Label |
biotin
|
| Label protocol |
RNA was processed in collaboration with Eurofins Genomics following standard Affymetrix protocol: Affymetrix IVT Pico Amp Kit, Ref 902762(30rxn)/902786(96rxn) was used to amplify 2ng input RNA. cRNA is then converted to biotinylated double-stranded cDNA (ds-cDNA) hybridization targets, all according to Affymetrix protocol.
|
| |
|
| Hybridization protocol |
RNA was processed in collaboration with Eurofins Genomics following standard Affymetrix protocol: Following fragmentation and biotin labelling of amplified ds cDNA, hybridization was performed using the Affymetrix Hyb., wash and stain kit (Ref 900720-C) and in hybridization oven for 16h at 45 °C. Chip type U133 Plus 2.0. GeneChips were washed and stained in the Affymetrix Fluidics Station 450.
|
| Scan protocol |
RNA was processed in collaboration with Eurofins Genomics following standard Affymetrix protocol: Arrays were scanned by Eurofins with a GeneChip Scanner 3000 (Affymetrix) according to manufacturer's protocols.
|
| Description |
Gene expression data from freshly FACS isolated cells, without culture
|
| Data processing |
RNA was processed in collaboration with Eurofins Genomics following standard Affymetrix protocol: The expression data was analyzed into Robust Multi-Array Average (RMA) normalized data by Eurofins.
|
| |
|
| Submission date |
Nov 17, 2022 |
| Last update date |
Dec 01, 2023 |
| Contact name |
Jolanda de Vries |
| E-mail(s) |
jolanda.devries@radboudumc.nl, anouk.becker@radboudumc.nl
|
| Organization name |
Radboud Institute for Molecular Life Sciences, Radboudumc
|
| Department |
Tumor Immunology Department
|
| Street address |
Geert grooteplein Zuid 28
|
| City |
Nijmegen |
| ZIP/Postal code |
6525GA |
| Country |
Netherlands |
| |
|
| Platform ID |
GPL570 |
| Series (1) |
| GSE218218 |
Expression data from primary human healthy donor-derived CD14- cDC2s, CD14+ cDC2s and monocytes |
|
