Use of fluorescence threshold triggering and high-speed flow cytometry for rare event detection

A simple rare event detection method utilizing dual-parameter flow cytometry is described, which allows quantitation of specific cellular events at the level of two cells in 10(7) total cells. Using a standard unmodified single laser flow cytometer sampling at a rate of 25,000 events/sec and a fluorescence discriminator, 10(7) total cells are processed in 7 min. The assay involves precise characterization of instrument flow rates to calculate total events processed by the cytometer rather than accumulate total events in computer memory. This method of detecting rare events is demonstrated by using a model system of breast cancer cells labeled with a metabolically activated dye and serially diluted into normal peripheral blood. Potential applications include validation of methods to detect minimum residual disease following myeloablative therapy, detection of any remaining tumor cells following purging methods, and validation of methods to detect circulating fetal cells in maternal blood.