Sweat the small stuff

COMMENTARY Viruses are obligate intracellular parasites. How viral agents enter and replicate in cells are critical issues because of impact on cellular function and because of the need to find ways to block infection at different stages. Intermediate steps in entry and replication are hard to study. Conventional physical means of purification can be complex and time consuming. Even for the most intensively studied viruses such as HIV, it is not clear exactly how the virus loses its envelope once inside the cell or how the viral genome is shuttled around in the cytoplasm (1). In this issue, Loret et al. (page 950) show elegant flow cytometry that they use to detect and sort Herpes simplex virus (HSV-1) type I nuclear particles based on differential DNA staining and side scatter (90 ). HSV-1 is a large double stranded DNA virus which causes cold sores in humans. The major nuclear particles of HSV-1 (A, B, C) were identified and sorted for biochemical analysis. The viral particles that have the highest signal after DNA staining with syto 13 were shown to scatter more light at 90 perhaps indicating that structural changes occur in the nucleus as the viral particles mature. This work represents an extension of cytometry into a new arena and offers new strategies for analysis and separation that can be used by virologists and other seekers of intracellular components. In addition, this paper provides important lessons for how best to do cytometry with small particles. The lessons learned include: