The purpose of this study was to examine how prior knowledge of cellular transport influenced how high school students in the USA viewed and interpreted graphic representations of this topic. The participants were Advanced Placement Biology students (n = 65); each participant had previously taken a biology course in high school. After assessing prior knowledge using the Diffusion and Osmosis Diagnostic Test, two graphical representations of cellular transport processes were selected for analysis. Three different methods of data collection--eye tracking, interviews, and questionnaires--were used to investigate differences in perceived salient features of the graphics, interpretations of the graphics, and processing difficulty experienced while attending to and interpreting the graphics. The results from the eye tracking data, interviews, and instructional representation questionnaires were triangulated and revealed differences in how high and low prior knowledge students attended to and interpreted particle differences, concentration gradient, the role of adenosine triphosphate , endocytosis and exocytosis, and text labels and captions. Without adequate domain knowledge, low prior knowledge students focused on the surface features of the graphics (ex. differences in particle colour) to build an understanding of the concepts represented. On the other hand, with more abundant and better-organised domain knowledge, high prior knowledge students were more likely to attend to the thematically relevant content in the graphics, which enhanced their understanding. The findings of this study offer a more complete understanding of how differentially prepared learners view and interpret graphics and have the potential to inform instructional design. (Contains 2 figures and 3 tables.)