PURPOSE: The increasing availability of fluorescent probes for in vivo optical imaging enables the interrogation of complex biological processes in small animals serving as models for human-like tissue function and disease. However, the validation of probe bio-distribution during their development or the study of different disease models, in support of in vivo imaging studies, is not straightforward. PROCEDURES: The imaging system developed consists of a customized multispectral planar imager that has been adapted on a commercial cryomicrotome and provides a powerful modality for ex vivo imaging of small animals. RESULTS: The ability to capture 3D anatomical (color) and fluorescence volumetric distributions of multiple fluorescent markers in high resolution is showcased. CONCLUSIONS: Serving both as a method for accurately imaging the bio-distribution of multiple fluorescent agents inside organisms and as a modality for the validation of non-invasive methods, multispectral cryoslicing imaging offers useful insights to ex vivo optical imaging of molecular probes.
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