Hypericin, a potent necrosis avid agent, features a peculiar affinity for necrotic tissue. Necrosis avid contrast agents have been investigated as markers for non-invasive imaging of different disorders. In view of the promising clinical applications, a more complete knowledge of the mechanism of action is important for the future development of new chemical structures with improved characteristics. To study whether a compound-specific or non-specific mechanism based on plasma lipoprotein transport is involved in the accumulation of hypericin in intratumoral necrosis, we performed a visual and quantitative fluoromicroscopic analysis of the colocalization of hypericin and DiOC18-labeled lipoproteins in subcutaneous murine radiation-induced fibrosarcoma tumors. Microscopic fluorescent overlay images of necrotic tumors demonstrated that hypericin already showed clear necrosis avid characteristics 4 h after injection, whereas a similar outstanding accumulation in necrosis was not demonstrated for the labeled lipoproteins. Moreover, a quantitative analysis of fluoromicroscopic images of tumor necrosis at 24 h after injection showed differences in normalized fluorescence intensities between hypericin and labeled lipoproteins of 50-100%, reflecting a shifted pattern in localization. We conclude that our results are indicative of a release of hypericin from the lipoprotein complex at some point along its way through the peri-necrotic tumor area and the necrotic tissue debris, which is in line with the hypothesis of a compound-specific mechanism.