Qualitative changes in activated T cells induced by checkpoint blockade in vivo

Here we perform QuantSeq 3' mRNA sequencing of RNA extracted from flow sorted splenic T cell from Tg4 Nr4a3-Tocky Tiger mice that had been immunised with 80ug of 4Y MBP peptide for 24 hours. After 24 hours mice were re-stimulated with 8ug 4Y MBP in the presence of isotype, anti-PD1 or anti-Lag3 antibodies for 4 hours. The results show that anti-PD1 but not anti-Lag3 imparts a unique transcriptional signature on responding T cells, indicative of stronger TCR signalling. Overall design: 3 experimental groups in biological duplicates. Tg4 Nr4a3-Tocky Tiger mice were immunised with 80ug of 4Y MBP peptide s.c. for 24 hours. After 24 hours, mice were injected i.p. with 800ug of either 1) Isotype, 2) anti-PD1(clone 29F.1A12) or 3) anti-Lag3 (clone CB97W) antibodies. 30 minutes later 8ug of 4Y MBP peptide was administered s.c. and 4hrs later spleens removed. CD4+ Nr4a3-TimerBlue+TimerRed+ T cells were flow sorted from spleens for analysis of RNA