characteristics.text: ERC000011
characteristics.characteristic: ENA-CHECKLIST
characteristics.text: 2023-07-31T00:17:54Z
characteristics.characteristic: ENA-FIRST-PUBLIC
characteristics.text: 2023-07-31T00:17:54Z
characteristics.characteristic: ENA-LAST-UPDATE
characteristics.text: SAMEA110671082
characteristics.characteristic: External Id
characteristics.text: Instituto de Medicina Molecular Joao Lobo Antunes
characteristics.characteristic: INSDC center name
characteristics.text: 2023-07-31T00:17:54Z
characteristics.characteristic: INSDC first public
characteristics.text: 2023-07-31T00:17:54Z
characteristics.characteristic: INSDC last update
characteristics.text: public
characteristics.characteristic: INSDC status
characteristics.text: ERS12768660
characteristics.characteristic: SRA accession
characteristics.text: E-MTAB-12085:Sample 6
characteristics.characteristic: Submitter Id
characteristics.text: 12
characteristics.unit: week
characteristics.characteristic: age
characteristics.text: ArrayExpress
characteristics.characteristic: broker name
characteristics.text: T follicular helper cell and T regulatory cell
characteristics.characteristic: cell_type
characteristics.text: house mouse
characteristics.characteristic: common name
characteristics.text: Protocols: Healthy female C57BL/6 Foxp3GFP reporter mice aged between 8 to 12 weeks were used. The mice were immunized subcutaneously in the footpad with ovalbumin (Ovalbumin EndoFit, Invivogen, Catalogue #vac-pova) either emulsified 1:1 (v:v) with incomplete Freund's adjuvant (IFA, Sigma-Aldrich, Catalogue #F5506), or entrapped with CpG in polymeric nanoparticles (NP-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 30 μg of NP-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 with the following monoclonal antibodies or reagents: anti-CD4 (RM4-4, Biolegend), anti-CD185/CXCR5 (2G8, BD Bioscience), and Streptavidin (Biolegend). DAPI (Biolegend) was used for dead-cell exclusion. Tfh and Treg cells were enriched by fluorescence-activated cell sorting (FACS) on a BD FACSAria cell sorter using the following gating strategy: Tfh cells as CD4+GFP(Foxp3)-CXCR5+ and Treg cells as CD4+GFP(Foxp3)+CXCR5-. For OVA/IFA immunization subset, Tfh cells were used. For OVA/NP-CpG immunization, both Tfh cells from OVA/NP-CpG immunization and naïve Treg cells from non-immunization were mixed in 1:1 ratio before loading into the 10X Chromium equipment. Single-cell libraries of FACS-sorted CXCR5+ Tfh from OVA/IFA and OVA/NP-CpG immunizations were generated using the 10x Genomics Chromium Single Cell 5' V(D)J reagents (10x Genomics; PN-1000006 and PN-1000020) according to the manufacturer's protocol. Single-cell libraries of FACS-sorted CXCR5+ Tfh from OVA/IFA and OVA/NP-CpG immunizations were generated using the 10x Genomics Chromium Single Cell 5' V(D)J reagents (10x Genomics; PN-1000006 and PN-1000020) according to the manufacturer's protocol.
characteristics.characteristic: description
characteristics.text: adult
characteristics.characteristic: developmental stage
characteristics.text: normal
characteristics.characteristic: disease
characteristics.text: Foxp3-GFP
characteristics.characteristic: genotype
characteristics.text: not applicable
characteristics.characteristic: isolate
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: female
characteristics.characteristic: sex
characteristics.text: C57BL/6
characteristics.characteristic: strain
characteristics.text: Sample 6
characteristics.characteristic: title