As technologies for high throughput and high content screening continue to evolve, new platforms for quantitative cellular imaging will play an increasingly important role in identifying and profiling lead compounds. To gain insight into the effects of a compound on cell morphology or intracellular events, it is necessary to have quality images and the ability to enumerate thousands of data points for statistical relevance. Imaging flow cytometry combines many of the features of flow cytometry, microscopy and imaging as well as a number of unique characteristics. The result is an instrument capable of highly quantitative analysis of cellular behaviors such as receptor internalization, phagocytosis, cell-cell communication, apoptosis and nuclear translocation. This promising new technology and unique type of flow cytometry provides enhanced capabilities for highly multiplexed assays. Here, we review the capabilities of the ImageStream imaging cytometer and discuss several applications relevant to compound screening and profiling.