The ability to monitor the spatial and temporal organization of molecules such as biopolymers within a cell is essential to enable the ability to understand the complexity and dynamics existing in biological processes. However, many limitations currently exist in specifically labeling proteins in living cells. In our study, we incorporate nanometer-sized semiconductor quantum dots (QDs) into living cells for spatiotemporal protein imaging of actin polymers in Dictyostelium discoideum without the necessity of using complicating transmembrane transport approaches. We first demonstrate cytoplasmic distribution of QDs within these living amoebae cells and then show molecular targeting through actin filament labeling. Also, we have developed a microfluidic system to control and visualize the spatiotemporal response of the cellular environment during cell motility, which allows us to demonstrate specific localization control of the QD-protein complexes in living cells. This study provides a valuable tool for the specific targeting and analysis of proteins within Dictyostelium without the encumbrance of transmembrane assisted methods, which has implication in fields including polymer physics, material science, engineering, and biology.