The utilization of self-assembly through non-covalent forces such as electrostatic interactions and hydrogen bonds offers an effective strategy for the construction of organic-inorganic intercalation composite materials. Although self-assembly strategies hold significant potential, the embedding of organic guest molecules within inorganic host materials through self-assembly processes remains unexplored. In this work, the real-time observation of the self-assembly process of organic-inorganic intercalation composite materials has been achieved using confocal microscopy. We found that methylene blue intercalated molybdenum oxide (MB-MoO3-x) can be synthesized using a mild solution-phase self-assembly reaction and proposed a potential crystal growth model. Our findings indicate that the intercalation of MB remarkably increases the interlayer spacing of MoO3, that causes a transition from an amorphous to a more ordered crystalline structure. Adjusting UV irradiation and N-Methyl-2-pyrrolidone (NMP) content, the particle size of MB-MoO3-x could be controlled, ranging from 400 nm to 500 μm. Characterization techniques such as XPS, FTIR and Raman spectroscopy revealed that the self-assembly of MB with MoO3 is driven by non-covalent electrostatic interactions. Our research provides significant insights into the self-assembly mechanism of organic-inorganic intercalation composite materials, aiding the development of materials that combine the stability and electronic utility of inorganic metal oxides with the chemical diversity and functionality of organic molecules.