This study compared a panel of three different prion protein antibodies with conventional plaque staining methods - Congo Red, Periodic acid Schiff and sulphated Alcian blue - to investigate amyloid plaque formation in cases of human spongiform encephalopathy (HSE). Tissue samples were taken from the cerebellum in nine sporadic cases of Creutzfeldt-Jakob Disease, with plaque formation noted on routine histology, and one case of Gerstmann-Straussler Scheinker syndrome. Using image analysis techniques, a semi-automatic system of plaque quantification was devised to measure the relative performance of these different staining methods. A total figure of percentage tissue area stained positively was returned by the system in each case analysed. A significant statistical correlation was observed among all three antibodies (r > 0.9, P < 0.01, in all comparisons) and a significant improvement was observed when the average antibody staining figures were compared to those of the Alcian blue technique (P < 0.05). The distribution of plaques across the cerebellar layers observed here appears to confirm earlier research findings. A strong correlation was found between staining in the two cerebellar hemispheres (r = 0.97, P < 0.01). This novel image analysis system has considerable potential for objective assessment of the pathology of HSE.