<p>[18F]-Fluoro-2-deoxyglucose positron emission tomography (FDG-PET) has been introduced as a method for response evaluation following neoadjuvant chemotherapy in pancreatic cancer. Glycolytic activity detected by FDG-PET is deemed to correlate with the residual cancer burden after treatment, but the expression of key glycolytic markers in cancer and stromal cells of pancreatic cancer after treatment remains unknown. The current study aimed to investigate the expression and inter-/intratumour heterogeneity of glucose transporter 1 (Glut1) and monocarboxylate transporter 4 (MCT4) in cancer cells and stroma of treatment-naïve and neoadjuvantly treated pancreatic cancer. Expression of Glut1 and MCT4 in cancer cells and stroma was investigated by immunohistochemistry in three tissue sections from 30 treatment-naïve and 30 neoadjuvantly treated pancreatic cancer tumours (total: 180 sections). An immunoreactive score was calculated as the percentage of positive cells multiplied with the intensity of labelling. The expression of glycolytic markers varied considerably between and within both treatment-naïve and neoadjuvantly treated tumours. In cancer cells, expression did not significantly differ between the groups, while in tumour stroma, the expression of glycolytic markers was significantly lower in neoadjuvantly treated compared to treatment-naïve tumours. Maintained expression of key glycolytic markers in cancer cells following neoadjuvant therapy supports the use of FDG-PET for response evaluation in pancreatic cancer. However, tumour stroma, which in some cases markedly expresses Glut1 and MCT4, may constitute a potential source of error. Clinical trials correlating FDG-PET with immunohistochemical assessment of glycolysis in treatment-naïve and neoadjuvantly treated pancreatic cancer are needed.</p>