Background and aims: Typically, an X-ray image forms as the applied ray becomes attenuated by the imaged sample. In biological and biomaterial samples, the common imaging issue is the low density and its homogeneous distribution. Without externally supplied heavy elements, studied features may not be visible at all, or the micro-computational tomography (µCT) imaging may take unreasonable amount of time resources. The goal of this study was to test and optimize different background and specific contrast enhancement methods for overcoming these imaging issues. Materials and methods: The used background contrasting methods were based on iodine and phosphotungstic acid. Each staining method were iterated until satisfactory contrasting results were gained for examining the three dimensional anatomy of adult zebra fish. The ethanol based iodine was also used for studying the structural dissimilarities between dystrophic and healthy rat retinas. Stem cell monolayer samples were specifically stained by antibodies which were used for localizing metallic silver in the immediate proximity of the examined antigens. Results: During the research it was shown that all of the used contrasting methods were capable for enhancing the studied features in the treated samples. All of the background contrasting methods successfully enhanced the anatomical features of the adult zebra fish. Furthermore, the ethanol based iodine contrasting preserved the general morphology of the rat eyes and made possible for distinguishing clear structural dissimilarities between dystrophic and control retinas. The specific contrasting method enhanced successfully the target actin and lamin antigens in the examined stem cell monolayer samples. Conclusions: The used contrasting methods gave good and comprehensive base for the following research for examining wide variety of biological samples with µCT.