Background: Cell behavior is the key to tissue regeneration. Given the fact that most of the cells used in tissue engineering are anchorage-dependent, their behavior including adhesion, growth, migration, matrix synthesis, and differentiation is related to the design of the scaffolds. Thus, characterization of the scaffolds is highly required. Micro-computed tomography (micro-CT) provides a powerful platform to analyze, visualize, and explore any portion of interest in the scaffold in a 3D fashion without cutting or destroying it with the benefit of almost no sample preparation need. Main body: This review highlights the relationship between the scaffold microstructure and cell behavior, and provides the basics of the micro-CT method. In this work, we also analyzed the original papers that were published in 2016 through a systematic search to address the need for specific improvements in the methods section of the papers including the amount of provided information from the obtained results.