Laboratory of Immunoregulation and Mucosal Immunology, VIB Center for Inflammation Research, Ghent, Belgium; Department of Internal Medicine and Pediatrics, Ghent University, Ghent, Belgium
Sahine Lameire
Laboratory of Immunoregulation and Mucosal Immunology, VIB Center for Inflammation Research, Ghent, Belgium; Department of Internal Medicine and Pediatrics, Ghent University, Ghent, Belgium
Manon Vanheerswynghels
Laboratory of Immunoregulation and Mucosal Immunology, VIB Center for Inflammation Research, Ghent, Belgium; Department of Internal Medicine and Pediatrics, Ghent University, Ghent, Belgium
Julie Deckers
Laboratory of Immunoregulation and Mucosal Immunology, VIB Center for Inflammation Research, Ghent, Belgium; Department of Internal Medicine and Pediatrics, Ghent University, Ghent, Belgium
Caroline De Wolf
Laboratory of Immunoregulation and Mucosal Immunology, VIB Center for Inflammation Research, Ghent, Belgium; Department of Internal Medicine and Pediatrics, Ghent University, Ghent, Belgium
Wendy Toussaint
Laboratory of Immunoregulation and Mucosal Immunology, VIB Center for Inflammation Research, Ghent, Belgium; Department of Internal Medicine and Pediatrics, Ghent University, Ghent, Belgium
Rein Verbeke
Laboratory of General Biochemistry and Physical Pharmacy, Faculty of Pharmaceutical Sciences, Ghent University, Ghent, Belgium
Kevin Verstaen
VIB Single Cell Core, VIB Center, Ghent, Belgium; Department of Applied Mathematics, Computer Science and Statistics, Ghent University, Ghent, Belgium
Hamida Hammad
Laboratory of Immunoregulation and Mucosal Immunology, VIB Center for Inflammation Research, Ghent, Belgium; Department of Internal Medicine and Pediatrics, Ghent University, Ghent, Belgium
Stijn Vanhee
Laboratory of Immunoregulation and Mucosal Immunology, VIB Center for Inflammation Research, Ghent, Belgium; Department of Internal Medicine and Pediatrics, Ghent University, Ghent, Belgium; Department of Head and Skin, Ghent University, Ghent, Belgium
Laboratory of Immunoregulation and Mucosal Immunology, VIB Center for Inflammation Research, Ghent, Belgium; Department of Internal Medicine and Pediatrics, Ghent University, Ghent, Belgium; Department of Pulmonary Medicine, Erasmus University Medical Center Rotterdam, Rotterdam, Netherlands
Since the precursor frequency of naive T cells is extremely low, investigating the early steps of antigen-specific T cell activation is challenging. To overcome this detection problem, adoptive transfer of a cohort of T cells purified from T cell receptor (TCR) transgenic donors has been extensively used but is not readily available for emerging pathogens. Constructing TCR transgenic mice from T cell hybridomas is a labor-intensive and sometimes erratic process, since the best clones are selected based on antigen-induced CD69 upregulation or IL-2 production in vitro, and TCR chains are polymerase chain reaction (PCR)-cloned into expression vectors. Here, we exploited the rapid advances in single-cell sequencing and TCR repertoire analysis to select the best clones without hybridoma selection, and generated CORSET8 mice (CORona Spike Epitope specific CD8 T cell), carrying a TCR specific for the Spike protein of SARS-CoV-2. Implementing newly created DALI software for TCR repertoire analysis in single-cell analysis enabled the rapid selection of the ideal responder CD8 T cell clone, based on antigen reactivity, proliferation, and immunophenotype in vivo. Identified TCR sequences were inserted as synthetic DNA into an expression vector and transgenic CORSET8 donor mice were created. After immunization with Spike/CpG-motifs, mRNA vaccination or SARS-CoV-2 infection, CORSET8 T cells strongly proliferated and showed signs of T cell activation. Thus, a combination of TCR repertoire analysis and scRNA immunophenotyping allowed rapid selection of antigen-specific TCR sequences that can be used to generate TCR transgenic mice.
