To compare regional variations in uptake of 3′-deoxy-3′- [ 18 F]-fluorothymidine (FLT) images using positron-emission tomography (PET) with measures of cellular proliferation from biopsy specimens obtained by image-guided brain biopsies.Fourteen patients with a supratentorial glioma that required an image-guided brain biopsy were imaged preoperatively with dynamic PET after the administration of FLT. Maps of FLT irreversible uptake rate (K i ) and standardized uptake value (SUV) were calculated. These maps were co-registered to a gadolinium-enhanced T1-weighted spoiled gradient echo (SPGR) sequence that was used for biopsy guidance, and the mean and maximum K i and SUV determined for each biopsy site. These values were correlated with the MIB-1 labelling index (a tissue marker of proliferation) from these biopsy sites.A total of 57 biopsy sites were studied. Although all measures correlated with MIB-1 labelling index, K i max provided the best correlation (Pearson coefficient, r=0.68; p<0.001). In low-grade gliomas the K i mean (±SD) was significantly higher than in normal tissue (3.3±1.7×10 −3 ml plasma /min/ml tissue versus 1.2±0.7×10 −3 ml plasma /min/ml tissue ; p=0.001). High-grade gliomas showed heterogeneous uptake with a mean K i of 7.7±4×10 −3 ml plasma /min/ml tissue . A threshold K i mean of 1.8×10 −3 differentiates between normal tissue and tumour (sensitivity 84%, specificity 88%); however, the latter threshold underestimated the extent of tumour in half the cases. SUV closely agreed with K i measurements.FLT PET is a useful marker of cellular proliferation that correlates with regional variation in cellular proliferation; however, it is unable to identify the margin of gliomas.