Project description:We tested orphan TCR autoreactivity using the peptide MHC-TCR chimeric receptor (MCR) co-culture system. In this system, cognate antigen recognition leads to TCR specific NFAT activation in MCR reporter cells expressing a mouse I-Ab MHC class II extracellular domain covalently linked to candidate peptides and an intracellular TCR signaling domain. We used mixed autoimmune bone marrow chimera spleens and kidneys as sources of cDNA to generate a transcriptome-wide library of natural autoantigen peptides . We cloned this cDNA-derived peptide (CDP) autoantigen library into the MCR retroviral backbone and transduced NFAT reporter cells to make a murine autoantigen MCR reporter library (MCR-Lib). We then used this library to screen orphan TCRs identified by scTCR-seq for autoreactivity.
Project description:We used protein arrays to measure IgG2c autoantibodies associated with Connective Tissue Diseases (CTDs) from retrogenic chimeras Sera was isolated from irradiated Icos-/-;CD45.1 mice reconstituted with 564Igi;Icos-/- bone marrow mixed with Icos-/-;CD45.1 bone marrow and WT bone marrow (BMchim.564) or retrogenic HSCs expressing TCR-A (BMchim.564-Icos.RAG-TCRA), TCR-B (BMchim.564-Icos.RAG-TCRB), or TCR-C (BMchim.564-Icos.RAG-TCRC). 564Igi (564homo) and 564Igi;Icos-/- (564homo.Icos) sera were included as controls.
Project description:We used protein arrays to measure IgG2a autoantibodies associated with Connective Tissue Diseases (CTDs) from retrogenic chimeras Sera was isolated from irradiated Icos-/-;CD45.1 mice reconstituted with 564Igi;Icos-/- bone marrow mixed with Icos-/-;CD45.1 bone marrow and WT bone marrow (BMchim.564) or retrogenic HSCs expressing TCR-A (BMchim.564-Icos.RAG-TCRA), TCR-B (BMchim.564-Icos.RAG-TCRB), or TCR-C (BMchim.564-Icos.RAG-TCRC). 564Igi (564homo) and 564Igi;Icos-/- (564homo.Icos) sera were included as controls.
Project description:We used protein arrays to measure IgG autoantibodies associated with Connective Tissue Diseases (CTDs) from retrogenic chimeras Sera was isolated from irradiated Icos-/-;CD45.1 mice reconstituted with 564Igi;Icos-/- bone marrow mixed with Icos-/-;CD45.1 bone marrow and WT bone marrow (BMchim.564) or retrogenic HSCs expressing TCR-A (BMchim.564-Icos.RAG-TCRA), TCR-B (BMchim.564-Icos.RAG-TCRB), or TCR-C (BMchim.564-Icos.RAG-TCRC). 564Igi (564homo) and 564Igi;Icos-/- (564homo.Icos) sera were included as controls.