There is increasing interest towards visualization of tissue level with deep penetration depth in bioscience and medical research, mesoscopic imaging exchanges resolution for penetration depth, which offers hundreds of micrometer resolution and up to several millimeter penetration depth. By introducing fluorescence dyes or with intrinsic fluorescence, much higher contrast of images can be recorded compared to reflection imaging. To assess the characteristics of fluorescence imaging system, in the first part of this thesis I will discuss 3D printing technique as a novel phantom fabrication method which enables the fabrication of optically realistic and morphologically complex tissue-simulating phantoms for the development and evaluation of optical imaging products. In the second part, I will discuss about the techniques and applications of Angled fluorescence laminar optical tomography (aFLOT), a modified fluorescence tomographic imaging technique based on 3D reconstructions. To extend the capability of aFLOT to acquire more bioinformation besides, its availability for quantification and statistics has been studied. Some technical improvements of aFLOT system performance has also been taken by incorporating algorithms for imaging processing. In addition, examples of biomedical applications have been discussed to demonstrate the capability of aFLOT system in both bioscience and medical research field.