Flow cytometry is the quickest and best way to measure fluorescently labeled properties of tens of thousands of individual cells in a single-cell suspension. A flow cytometer counts thousands of cells per second and does it objectively, instead of relying on the "speed" and subjectivity of someone laboriously peering through a fluorescent microscope. Spectrofluorometers aren't much better as they only measure the average property of a large number of cells, but flow cytometers measure each cell individually, which can show whether or not a sample is behaving heterogeneously. Further, by measuring several properties of each cell simultaneously, the flow cytometer can determine which population of a heterogeneous sample is behaving differently. Interesting populations can even be viably sorted to 99% purity for further studies or cloning experiments - from a single cell in one well of a multi-well culture plate to millions of cells in a test tube - impossible with an image cytometer.
Flow cytometry is quite flexible, and can be used in a variety of applications. The fluorescent dye, or fluorochrome, may bind directly to the property to be measured, including: DNA (cell cycle analysis), RNA, total/basic protein, surface sugars/lectins, thiols/glutathione, and lipids. Fluorochromes can also be conjugated to monoclonal antibodies to detect cell surface and intracellular antigens. Some fluorochromes detect functional properties of cells, for example: viability, enzyme activity, intracellular receptors, or DNA synthesis (cell kinetics). Cell activation can also be measured by: cell surface ligand binding, cell membrane potential, mitochondrial membrane potential, intracellular calcium, and intracellular pH. Some more recent applications in flow cytometry include apoptosis measurements and use of green fluorescent protein as a gene reporter.
-- Eric Van Buren, Manager, Flow Cytometry Core Facility
