Chapter 8 - Principles and applications of flow cytometry

Abstract

Flow cytometry techniques are widely employed to identify phenotypes and to explore mechanisms in cell-based assays. The principle of cytometry involves passage of cells through a light beam in a capillary where detectors and filters are positioned to capture signaling. These detectors measure absorbed or scattered light by the sample, making possible a morphological analysis. In addition, the use of fluorescent markers extends the possibilities of using the equipment. The genetic material of cells can be labeled with fluorophores to evaluate the cell cycle. Cells can be externally stained for proliferation analysis. Extracellular, intracellular, or even intranuclear proteins can also be labeled, making it possible to identify and characterize phenotypes. Secreted proteins can also be complexed to appropriate beads and labeled with fluorescent antibodies, allowing their quantification. In addition to enabling the identification and characterization of cells, specialized equipment also has a cell-sorting feature that allows the separation of cell types for later cultivation and application. More sophisticated devices already allow for simultaneous analysis, such as fluorescence microscopy and single cell sequencing. In this chapter, we will discuss some concepts and applications of flow cytometry, as a powerful analytical tool for cell biology laboratories.