characteristics.text: ERC000011
characteristics.characteristic: ENA-CHECKLIST
characteristics.text: 2023-03-16T00:17:37Z
characteristics.characteristic: ENA-FIRST-PUBLIC
characteristics.text: 2023-03-16T00:17:37Z
characteristics.characteristic: ENA-LAST-UPDATE
characteristics.text: SAMEA10838336
characteristics.characteristic: External Id
characteristics.text: AstraZeneca R&D
characteristics.characteristic: INSDC center name
characteristics.text: 2023-03-16T00:17:37Z
characteristics.characteristic: INSDC first public
characteristics.text: 2023-03-16T00:17:37Z
characteristics.characteristic: INSDC last update
characteristics.text: public
characteristics.characteristic: INSDC status
characteristics.text: ERS8489570
characteristics.characteristic: SRA accession
characteristics.text: E-MTAB-11116:X07_S7_tet
characteristics.characteristic: Submitter Id
characteristics.text: 6 to 8
characteristics.unit: week
characteristics.characteristic: age
characteristics.text: ArrayExpress
characteristics.characteristic: broker name
characteristics.text: CD8-positive, alpha-beta T cell
characteristics.characteristic: cell type
characteristics.text: house mouse
characteristics.characteristic: common name
characteristics.text: Protocols: Single cell suspension was prepared from lungs using the mouse lung dissociation kit (Miltenyi Biotec) and passed through a 70µm strainer. Cells were labelled with fluorochrome conjugated antibodies and different cell populations were sorted in RLTplus buffer (Qiagen) using BD FACS ARIAIII and FACS DIVA software (BD Biosciences). For sorting of tetramer (NP) positive cells, single cell suspensions from lungs were enriched for CD8+ T cells using EasySep™ Mouse CD8+ T Cell Isolation Kit (Stem Cell technologies) prior to staining with antibodies. 6-8 weeks old Balb/cJRj mice (Janvier Labs) were subjected to intranasal (i.n.) sensitization/challenge regimen over a span of three weeks using house dust mite (HDM) extract from Dermatophagoides pteronyssinus (Stallergenes Greer, USA). In the first and third week, the mice were sensitized/challenged with 25mg of HDM extract in 20ml of phosphate-buffered saline (PBS) daily for 5 consecutive days. 21 days after the last HDM dose, the mice were immunized once with a sublethal dose of X47 H3N2 flu virus (0.1LD50, 1 PFU in 30ml of PBS) or PBS only as control via the i.n. route. Control groups of mice that were only sensitized/challenged with HDM, only immunized with X47 H3N2 flu virus or PBS-treated controls were also included. Seven weeks after the immunization with X47 (day 87), mice were sacrificed to study influenza- and HDM-specific memory immune responses. Total RNA was extracted from sorted T cell population using RNeasy plus micro kit (Qiagen) following manufacturer’s instructions Libraries were generated using QIAseq FX single cell RNA library kit (Qiagen) according to manufacturer’s instructions. Libraries were validated on the Fragment Analyzer platform (AATI) and concentrations were determined using the Quant-iT dsDNA HS assay kit on the Qubit fluorometer (Thermo Fisher Scientific) as well as with qPCR using QIAseq Library Quant Assay kit.
characteristics.characteristic: description
characteristics.text: adult
characteristics.characteristic: developmental stage
characteristics.text: Influenza A virus (A/X-47(H3N2))
characteristics.characteristic: infect
characteristics.text: Mus musculus
characteristics.characteristic: organism
characteristics.text: Mus musculus
characteristics.characteristic: organism
characteristics.text: lung
characteristics.characteristic: organism part
characteristics.text: tetramer (NP) positive
characteristics.characteristic: phenotype
characteristics.text: Mus musculus
characteristics.characteristic: scientific name
characteristics.text: female
characteristics.characteristic: sex
characteristics.text: PBS
characteristics.characteristic: stimulus
characteristics.text: Balb/cJRj
characteristics.characteristic: strain
characteristics.text: X07_S7_tet
characteristics.characteristic: title