Identification of HLA-DR as an antigen for a Vdelta1-positive gamma-delta TCR

We aimed to identify new ligands for Vdelta1-positive gamma-delta TCRs. To this end, we performed reporter cell screenings and identified Vgamma3 Vdelta1 positive gamma-delta TCR recognizing Raji B cell lymphoma cells. With soluble ectodomains of the respective TCR binding to Raji cells, we were able to further perform a genome-wide CRISPR-Cas9 knock-out screening and selected cells that lost binding to soluble version of the gamma-delta TCR of interest. The contained gRNAs were amplified via PCR and sequenced via Illumina NGS. This led to the identification of HLA-DR as the cognate antigen. Via gene-specific PCR amplification and subsequent Illlumina NGS, we identified the HLA-DR haplotype of Raji cells.The used Human Brunello CRISPR knockout pooled library was a gift from David Root and John Doench (Addgene #73178).