Diffuse optical tomography (DOT) employs near infrared (NIR) diffused light to probe functional information of biological tissues. To further increase a target to background contrast, fluorescent diffuse optical tomography (FDOT) has been proposed in recent years. Assisted by proper molecular probes, FDOT has been expanded to perform molecular imaging in small animals with high sensitivity and specificity. 

Unfortunately, conventional FDOT suffered from low reconstruction accuracy because the number of optical measurements is much less than the number of unknowns to be reconstructed. In this study, we developed a new reconstruction technique to improve the reconstruction accuracy by properly reducing the unknown number. Unlike conventional FDOT, we separated the information of a target into two categories: structural and functional parameters. The central position, depth and size of the target were considered as the structural parameters. The fluorophore concentration and lifetime were viewed as the functional parameters. The reconstruction process was divided into two steps. In the first step, ratios between different raw data were used to reconstruct the structural parameters. Once the structural parameters were recovered, the imaging position and volume were localized around the target region, which greatly reduced the unknown number. Based on the localization in the first step, the raw data, rather than the ratios of the data, were used to reconstruct the functional parameters in the second step. Experiments showed that this technique provided more accurate reconstruction results for both the structural and the functional parameters of the target than the conventional techniques.

Another disadvantage of conventional FDOT is the low spatial resolution. In order to improve the resolution, we proposed a novel fluorescence imaging technique. In this technique, a focused ultrasound beam was proposed to modulate the fluorophore concentration only within the focal zone of the ultrasound beam. Based on a Rate equation with a two-energy model, we found that the modulated fluorophore concentration were converted into modulated fluorescence emission signals that could be detected and used to extract the concentration and lifetime of the fluorophore in the focal zone of the ultrasound beam. By scanning the sample or the measurement system, a spatial distribution of fluorophore concentration or lifetime could be obtained. The spatial resolution of this hybrid technique depends only on the ultrasound resolution that is much higher than the resolution of the diffuse optical imaging techniques.