HeLa cells were infected with Yersinia pseudotuberculosis for 0.5-3 h. Intracellular bacteria could then be demonstrated by three different techniques: viable count, fluorescent-antibody staining and electron microscopy. Most of the bacteria seemed to be viable, since there was a good positive correlation (0.94) between viable and fluorescent bacteria. The bacterial uptake seemed to be mediated by a phagocytic-like procedure. The intracellular bacteria seemed to reside in vacuoles some of which increased in size as a function of time. The kinetics of infection was studied after addition of 10(7) or 10(9) bacteria per cell culture (2 X 10(6) cells). After a lag period of about 30 min there was a linear increase of intracellular bacteria, and this uptake proceeded for 1-2 h until most of the bacteria were ingested or an upper limit of ingested bacteria was reached. The upper limit was calculated to be a mean of 60 per infected cell in the cell culture. More than 90% of the cells could be infected and a reasonable number of the bacteria survive in the cells for at least 3 days, as demonstrated by the viable-count technique. The bacteria-cell system may be used to study, for example, the effect of antibiotics or antibodies on intracellular bacteria and pathogenicity of intracellular diseases.