Objectives Spectral imaging as the latest undestroyed identification method is being paid more and more attention by forensic science researchers ever since its introduction into the field. The general law of the selection of region of interest in spectral imaging technique makes the main purpose this paper tries to solve via experiments. In this study, spectral images were classified and analyzed under different regions of interest by specialist spectral imaging software. Tt was found that there are much more spectrum differences among various concentration regions even if the specimen is uniform. It’s clearly that selecting different regions of interest is quite important for forensic analysis and classification. It is summarized that imaging quality is affected by the selection of regions of interest and the concentration in visible scope.