What Are the Principles of Flow Cytometry?

Andrew Kirmayer
Andrew Kirmayer

Flow cytometry is often used to determine the nature of individual cells that flow, one by one, past a laser. It can determine the size of cells, the components of their cytoplasm, the state of genetic material, and the presence of proteins in the cell and on the membrane. The optical and fluorescent properties of cells can also be measured. In many scientific and medical disciplines, the principles of flow cytometry typically include labeling cells with fluorescent dyes, and exciting them with a light source such as a laser. Particles are generally stimulated into a higher energy state and often emit light when they return to a resting state; measurements of color and wavelength can allow many properties of cells to be measured.

One of the principles of flow cytometry involves how cells are labeled. Dyes such as propidium iodide, flouorescein, and phycoerythrin are often used, and can be combined for measuring more than one characteristic to perform multi-parametric analysis. Cells are generally drawn through the instrument into a fluid. The mixture then usually flows past a light source while specialized optics direct the emitted light into the device’s mirrors and filters. Optical detectors can convert the light signals into electrical pulses; this is often accomplished with a photomultiplier tube.

An analog-to-digital converter sometimes amplifies the signals which can then be sent to a computer to be graphed. The results can be analyzed using one-parameter histograms that graph cell count versus a specific measurement; graph height typically corresponds to the voltage generated by the light emission. A two-parameter histogram is often used to measure cell count related to density. Some instruments can provide data related to different levels of light scattering, while others expand on the principles of flow cytometry with the capability of sorting cells based on separate classifications of data.

Flow cytometry can be used to study genetics, cell cycles, and various aspects of biology. The principles of flow cytometry have often been applied for diagnostics, including the analysis of deoxyribonucleic acid (DNA) and abnormalities with it that can be associated with blood cancers, for example. Clinical scientists can often provide insight into a prognosis based on the medical condition and DNA analysis.

The electronic detection of various immune cells can be performed as well. Using the principles of flow cytometry, blood, bone marrow, and solid tissue samples can be analyzed. Other body fluids, such as urine and cerebrospinal fluid, are also often studied in this manner. Various software programs can collect and graphically display data as samples are run through an instrument, which is often small and inexpensive enough for use in basic medical clinics.

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