Glycosylation changes on surface molecules of T cells affect cell trafficking and function and may be useful in discriminating between naive, effector, and memory T cells. To analyze oligosaccharide structures on T cells activated in vivo, we examined alterations in sialic acid residues on T cells following infection of mice with lymphocytic choriomeningitis (LCMV), vaccinia virus, and vesicular stomatitis virus. We found that the majority of CD8 T cells from mice acutely infected with these viruses showed increased binding to peanut agglutinin (PNA). All of the PNAhighCD8 T cells from infected mice were CD44high, indicating that glycosylation changes were occurring on activated T cells. There was also an increase in the PNAhighCD4 T cell population in virally infected mice. Increased PNA binding to activated CD8 T cells correlated with higher endogenous neuraminidase levels in these cells. This higher neuraminidase activity most likely contributed to the PNAhigh phenotype by cleaving sialic acid residues off the core-1 O-glycans or glycoproteins destined for the cell surface. A PNAhighCD8 T cell population persisted in immune mice that had cleared the LCMV infection. When spleen cells from immune mice were sorted into PNAhigh and PNAlow populations, >95% of the LCMV-specific memory CD8 T cells segregated with the PNAhigh population. This shows that virus-specific memory CD8 T cells remain hyposialylated and can be distinguished from naive CD8 T cells based on PNA binding. Thus, PNA can be used as a marker for Ag-experienced T cells.