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Introduction to Flow Cytometry

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A flow cytometer consists of several component systems; fluidics pumps, lasers, fluorescence detectors, signal converters and computers. When combined, these systems can be used to analyze the physical characteristics of particles or cells in fluid suspension and sort them from mixed samples.

When samples are placed on a flow cytometer, particles are taken up in a solution that carries them into a flow cell where they pass by a laser one at a time. As the cells move by the laser, intrinsic and extrinsic properties are measured and then tramsitted to a computer display.

Measurable intrinsic cellular properties include relative cell size and complexity. These characteristics are based on the light scattering properties of membranes both inside and outside of the cell and provide the basis for selecting a population of cells from a non-uniform mixture for further analysis.

Determinations of other cellular properties are based on measures of fluorescence. Molecules on the cell can be specifically labeled with antibodies which are tagged with various light emitting molecules (fluorochromes). The presence of these labels on the cells is revealed after their tags are excited by laser light. These tags emit light (fluoresce) that is detected by photomultiplier tubes (PMT’s) at various wavelengths.

Tagging cells with a variety of antibodies bound to different colour emitting fluorocrhomes can reveal a lot about how cells respond to differnte stimuli. That is, the relative intensity of the fluoresce measured on cells can be related to cellular responses after known treatments or can be used to assess the signs of disease progression in complex immunological disorders such as HIV and autoimmunity.