Characterization of Flow Cytometer Instrument Sensitivity

Fluorescence sensitivity, measured in terms of resolution, allows the researcher to more accurately answer the question of how dim a cell can be and still be resolvable from another population. This measure focuses on the width, i.e., the standard deviation, of the population distributions and not on the location, i.e., the mean intensity, of populations on a histogram scale relative to the background. To determine the fluorescence sensitivity in terms of the resolution, both the detection efficiency, Q, and optical background, B, need to be measured. Both factors affect the ability to resolve dimly fluorescent subpopulations, and both factors are required to uniquely characterize the performance of a flow cytometer. Using Q and B, it is possible to determine the minimum fluorescence intensity that is resolvable from the background or from another population. This unit describes a practical and robust approach to measure the critical factors that determine the ability of a flow cytometer to analyze dimly fluorescent particles. Keywords: fluorescence; sensitivity; resolution; Q; detection efficiency; background