Building and Optimizing a Virus-specific T Cell Receptor Library for Targeted Immunotherapy in Viral Infections

Banu, Nasirah; Chia, Adeline; Ho, Zi Zong; Garcia, Alfonso Tan; Paravasivam, Komathi; Grotenbreg, Gijsbert M.; Bertoletti, Antonio; Gehring, Adam J.

Next, we analyzed IFN-c production in Vbeta1 T cells. The advantage of Vbeta staining is that nearly all T cells expressing our introduced TCR, whether is it properly paired with the correct alpha chain or not, will be stained by the antibody. Therefore, Vbeta staining may detect T cells producing IFN-c that do not express enough of the introduced TCR for HLA-pentamer staining. Similar to pentamer staining, TCR beta chain expression was similar across all conditions (Fig. 6D ) and the frequency of IFN-c1 T cells increased with the addition of polyI:C and ssRNA40 (Fig. 6E) . Unlike T cells stained with pentamers, we found that nearly all (<90%) of the IFN-c1 cells following peptide-specific activation were TCR V beta positive (Fig. 6E&F) . These data indicate that IFN-c production was specific to the introduced TCR. They also confirm increased Th1 polarization as cells shift from Vbeta1/IFN-c-to Vbeta1/IFN-c1 with the addition of TLR ligands to the culture. Therefore, similar to their role in natural infection, combining T cell activation in the presence of TLR ligation led to an inflammatory environment during the activation phase for transduction and translated into a greater frequency of IFN-c producing T cells.