The aim of the present study was to investigate the content of organochlorine pesticides (OCPs) and polychlorinated biphenyls (PCBs) in muscle tissues of fish, i.e. carp (Cyprinus carpio, L.), bream (Abramis brama, L.) and pike perch (Stizostedion lucioperca, L.) from the middle Nové Mlýny reservoir and compare our results with previous corresponding studies. Samples were analysed by gas chromatography with ion trap tandem mass spectrometry. The highest contents of all pollutants were determined in muscle tissue of bream. The analysis of HCHs showed that β-HCH was the most abundant. PCB congener 28 was evaluated as the predominant PCB congener. The metabolite p,p'-DDE exhibited the highest concentration of all the monitored metabolites. The monitoring confirmed significantly (P < 0.01) higher concentrations of OCP and PCB in muscle of bream compared to carp and pike perch. The contents of pollutants in our study were found to be lower in comparison to the findings of some previous studies.