Abstract. Accurate tumor excision is crucial in the locoregional treatment of cancer, and for this purpose, surgeons often rely on guide wires or radioactive markers for guidance toward the lesion. Further improvement may be obtained by adding optical guidance to currently used methods, in the form of intra-operative fluorescence imaging. To achieve such a multimodal approach, we have generated markers that can be used in a pre-, intra-, and post-operative setting, based on a cocktail of a dual-emissive inorganic dye, lipids, and pertechnetate. Phantom experiments demonstrate that these seeds can be placed accurately around a surrogate tumor using ultrasound. Three-dimensional bracketing provides delineation of the entire lesion. Combined with the multimodal nature, this provides the opportunity to predetermine the resection margins by validating the placement accuracy using multiple imaging modalities ͑namely, x ray, MRI, SPECT/CT, and ultrasound͒. The dual-emissive fluorescent properties of the dye provide the unique opportunity to intra-operatively estimate the depth of the seed in the tissue via multispectral imaging: emission green max = 520 nmഛ 5 mm penetration versus emission red max = 660 nmഛ 12 mm penetration. By using particles with different colors, the original geographic orientation of the excised tissue can be determined.