(a,b) HSM-AD images of sub-dermal U87MG tumor xenografts from mice injected with LGNRs display distinct accumulation patterns depending on the molecular specificity of the LGNR surface coating. Anti-αVβ3 LGNRs exhibit 7.5-fold greater accumulation in tumor tissue (a) than spectrally-identical LGNRs with non-specific IgG antibody coating (b) (n = 4 FOVs for Anti-αVβ3 LGNRs, n = 5 FOVs for IgG-LGNRs,two-tailed Student’s t-test, p=0.0041). The greater uptake of anti-αVβ3 LGNRs may result in part from specific LGNR binding to αVβ3 integrin, which is over-expressed by U87MG cells. (c–f) Validation of HSM-AD images with dark-field images of slightly higher spatial resolution further indicates that a substantial portion of anti-αVβ3 LGNRs are located along the edges of small capillaries within the tumor tissue (c,e) while no such association is observed for IgG-LGNRs (d,f). This observation is consistent with the nature of angiogenic tumor vasculature, which is also characterized by high expression levels of αVβ3 integrin in the vascular endothelium. Individual erythrocytes within angiogenic capillaries are denoted by white arrows, and capillary edges are approximately outlined by dashed blue ovals (e,f). Discrete regions of anti-αVβ3 LGNRs were also observed outside of the tumor vasculature, presumably due to either (1) specific binding to αVβ3integrin expressed by U87MG cells and/or (2) non-specific accumulation via the enhanced permeability and retention (EPR) effect characteristic of tumors. The absence of IgG-LGNR extravascular accumulation suggests the former of these mechanisms as the predominant source of anti-αVβ3 LGNR uptake in tumor tissue.