Purpose C-11-acetate (C-11-ACE) uptake on PET/CT was recently discovered to represent reactive astrocytes in the tumor microenvironment. This study aimed at evaluating the role of C-11-ACE PET/CT as an imaging biomarker of reactive astrogliosis in characterizing different types of gliomas. Methods In this prospective study, a total of 182 patients underwent C-11-ACE PET/CT before surgery. The ratio of SUVmax of a glioma to the SUVmean of the contralateral choroid plexus (C-11-ACE TCR) on PET/CT was calculated. C-11-ACE TCRs were compared with the World Health Organization grades and isocitrate dehydrogenase 1 (IDH1) mutation status. Grade 2 was considered low-grade tumor, and grades 3 and 4 were considered high-grade tumors. Results The median C-11-ACE TCR was significantly higher in IDH1 wild-type (wt) tumors (n = 91) than in IDH1-mutant (mt) tumors (n = 91) (2.38 vs 1.30, P < 0.001). Of the 91 IDH1-mt tumors, there were no differences in the median C-11-ACE TCRs between oligodendrogliomas (ODs) and astrocytic tumors (1.40 vs 1.20, P > 0.05). In grading low- versus high-grade gliomas, the receiver operating characteristic curve analyses showed a higher area under the curve (0.951) in IDH1-wt tumors than in IDH1-mt tumors (0.783, P = 0.002). Grade 2 ODs were well differentiated from high-grade gliomas. The C-11-ACE TCR of grade 3 ODs was significantly lower than that of IDH1-wt glioblastomas. Conclusions High C-11-ACE uptake is associated with high-grade IDH1-wt tumors, thus facilitating differentiation from high-grade IDH1-mt and low-grade gliomas. In particular, low C-11-ACE uptake in ODs is advantageous in overcoming the limitation of radiolabeled amino acid tracers.