Since its introduction over 25 years ago, flow cytometry has evolved to be one of the most important methods in cellular diagnostics and research. Within recent years, new technology has dramatically expanded the range of parameters that can be analyzed by this technology. These include, for example, the production or secretion of soluble mediators (1-4 ), cell proliferation (5-8), and the detection of molecules expressed at very low density (9,10). One of the most prominent recent developments, however, is the analysis of antigen-specific T cells. The problems encountered when analyzing antigen-specific T cells are manyfold. One of the major problems is the fact that their frequencies tend to be rather low. Recent data show that, in certain situations (e.g., acute immune responses against viruses or bacteria), the actual frequency of specific CD8 cells can be much higher than previously estimated by limiting dilution assays; however, in most other responses, the frequencies of specific T cells are clearly below these values. Frequencies as low as 1CH-10—⁵ have been reported (11,12).