characteristics.text: ERC000011
characteristics.characteristic: ENA-CHECKLIST
characteristics.text: 2023-04-15T00:20:02Z
characteristics.characteristic: ENA-FIRST-PUBLIC
characteristics.text: 2023-04-15T00:20:02Z
characteristics.characteristic: ENA-LAST-UPDATE
characteristics.text: SAMEA110654284
characteristics.characteristic: External Id
characteristics.text: Instituto de Medicina Molecular Joao Lobo Antunes
characteristics.characteristic: INSDC center name
characteristics.text: 2023-04-15T00:20:02Z
characteristics.characteristic: INSDC first public
characteristics.text: 2023-04-15T00:20:02Z
characteristics.characteristic: INSDC last update
characteristics.text: public
characteristics.characteristic: INSDC status
characteristics.text: ERS12751866
characteristics.characteristic: SRA accession
characteristics.text: E-MTAB-12086:Sample 50
characteristics.characteristic: Submitter Id
characteristics.text: 8 to 12
characteristics.unit: week
characteristics.characteristic: age
characteristics.text: ArrayExpress
characteristics.characteristic: broker name
characteristics.text: CD4-positive helper T cell
characteristics.characteristic: cell_type
characteristics.text: house mouse
characteristics.characteristic: common name
characteristics.text: Protocols: Mice aged between 8 to 12 weeks were immunized subcutaneously in the footpad with ovalbumin (Ovalbumin EndoFit, Invivogen, #vac-pova) either emulsified 1:1 (v:v) with incomplete Freund's adjuvant (IFA, Sigma-Aldrich, #F5506), admixed with CpG (ODN 1826 - TLR9 ligand, Invivogen, #tlrl-1826), or entrapped with CpG in polymeric nanoparticles (nano-CpG). Each animal was inoculated in the paw with one of the antigen/adjuvant mixtures, with a volume of 50 μL per paw containing 80 μg of OVA and, in the case of CpG or NP-CpG formulations, 30 μg of CpG. Popliteal lymph nodes were collected on day 11 following immunization. Single-cell suspensions were obtained by disrupting the lymph nodes in PBS (Lonza) with 2% FBS (Gibco) with curved forceps and a nylon mesh. Surface stainings were performed in the same buffer and a viability dye was used for dead-cell exclusion. Cells were sorted on a BD FACSAria cell sorter directly into DNA LoBind tubes (Eppendorf, #0030108051) containing nuclease-free water (Sigma-Aldrich, #W4502) with 0.2% Triton X-100 (Sigma-Aldrich, #T8787) and RNAse inhibitor at 2 U/mL (RNaseOUT, Invitrogen, #10777019). Cells were frozen in dry ice and kept at -80 ºC until further processing. There was no RNA isolation, the cDNA was prepared directly from the cell lysate. cDNA preparation was done directly from cell lysatesat GeneCore EMBL facility, according to the protocol described by Picelli et al. for Smart-seq2 (Nat Protoc. 2014 Jan;9(1):171-81. doi: 10.1038/nprot.2014.006), and sequencing libraries were prepared based on the tagmentation protocol described by Hennig et al. (G3 (Bethesda). 2018 Jan 4;8(1):79-89. doi: 10.1534/g3.117.300257).
characteristics.characteristic: description
characteristics.text: adult
characteristics.characteristic: developmental stage
characteristics.text: Thy1.1 x Thy1.2
characteristics.characteristic: genotype
characteristics.text: Mus musculus
characteristics.characteristic: organism
characteristics.text: Mus musculus
characteristics.characteristic: organism
characteristics.text: lymph node
characteristics.characteristic: organism part
characteristics.text: Mus musculus
characteristics.characteristic: scientific_name
characteristics.text: male
characteristics.characteristic: sex
characteristics.text: C57BL/6
characteristics.characteristic: strain
characteristics.text: Sample 50
characteristics.characteristic: title