Flow Cytometric Analysis of Cell Surface Antigen Density

Analysis of cell surface antigen density and, thus, the distribution of cell surface molecules among individual members of complex cell populations can be achieved rapidly and accurately by flow cytometry. This approach has been revolutionized over the last 20 yr by the codevelopment of monoclonal antibodies (MAb) to an ever-wider selection of cell-associated molecules, novel fluorochromes, and flow cytometric technology backed by increasingly powerful computers. This combination of technologies allows the simultaneous measurement of independent fluorochrome markers as well as light scatter parameters of individual cells as they pass through an intense focused light source. The multiple analysis of individual cells enabled by flow cytometry has facilitated the definition of functional subpopulations of many cell types, and some aspects of this approach are discussed in Chapter 34. In this article, the potential of flow cytometric analysis for quantitative purposes is considered, exemplified by discussion of factors important for the measurement of antigen density by quantitation of antibody binding.