Direct measurement of neurotransmitter in a single cell still remains a major challenge. Existing probes have several problems for accurate observation of intracellular component. In recent years, quantum dots (Qdot), with their unique physical, chemical, and optical properties, have been used extensively as probes to visualize several cell membrane receptors and extracellular biomolecules. However, high affinity between quantum dots may induce serious aggregation in the cytoplasm; as a result, quantum dot aggregates are usually misinterpreted as quantum dot-probed intracellular molecules. Moreover, higher viscosity of cytoplasm compared to the extracellular aqueous media may lead to a higher number of aggregated quantum dots. To eliminate these obstacles, we developed a direct nondestructive serotonin imaging in an intact cell using the quantum dot-based immunoassay with a rapid tunable multicolor imaging system based on the acousto-optic tunable filter. It provides rapid image scanning of serotonin granules as a function of wavelength under conditions in which the serotonin granules are bound with different colored quantum dots anti-serotonin antibody conjugates. This multicolor imaging technique clearly does away with aggregation issues that emerge from the use of quantum dots as probes for intracellular molecules. The results of this work demonstrate that multicolor imaging combined with fluorescent semiconductor nanocrystals has the potential to serve as a viable technique for nondestructive intracellular serotonin measurements in intact cells.