<p>Grazing behavior of free-living aquatic heterotrophic nanoflagellates (HNFs) on bacteria plays a central role in shaping microbial community structure and driving nutrient cycling. However, direct observation of these predator–prey interactions has been limited by the rapid motility of flagellates and the transient nature of their encounters. To overcome these challenges, this study presents a novel application of video-rate two-photon fluorescence microscopy for high-resolution, real-time imaging of fast-moving microorganisms. Using the HNF <i>Cafeteria roenbergensis</i> as a model system, we investigate dynamic grazing interactions between fluorescently stained bacteria and the flagellates detected via their intrinsic cellular autofluorescence. This two-photon microscope combined with real-time imaging capability enables continuous observation of the full grazing sequence: contact, capture, ingestion, and digestion, at single-cell resolution. Quantitative analyses across varying prey concentration reveal phase-specific durations and saturation behavior in grazing activities. Furthermore, real-time tracking uncovers a previously unobserved transition in grazing dynamics across two feeding behaviors of flagellates from starved to fed states in motile flagellates. This technique provides a powerful new tool to study rapid microbial interactions in situ and can be broadly applicable to diverse microbe-microbe systems. With the integration of targeted fluorescent molecular probes, this technique offers significant potential to elucidate mechanical and biochemical processes underlying microbial feeding and communication.</p>