Pre-clinical monitoring of tumor growth and identification of distal metastasis requires a balance between accuracy and expediency. Bioluminescence imaging (BLI) is often used to track tumor growth but is primarily limited to planar 2-dimensional (2D) imaging. Consistent subject placement within a standard top-mounted, single-detector small animal imager is vital to reducing variability in repeated same-animal measures over time. Here, we describe a method for tracking tumor development using a multi-angle BLI and photo-acoustic workflow. We correlate serial caliper measurements and 2D BLI to 360° BLI and photo-acoustic datasets for the same animals. Full 360° BLI showed improved correlations with both volumes obtained from caliper measurements and photo-acoustic segmentation, as compared to planar BLI. We also determined segmented tumor volumes from photo-acoustic datasets more accurately reflects true excised tumors’ volumes compared to caliper measurements. Our results demonstrate the distinct advantages of both 360° surface mapping by BLI and photo-acoustic methodologies for non-invasive tracking of tumor growth in pre-clinical academic settings. Furthermore, our design is fully implementable in all top-mounted, single-detector imagers, thereby providing the opportunity to shift the paradigm away from planar BLI into rapid BLI tomography applications.

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