How Fluorescent Dye Is Used In Flow Cytometry

Flow cytometry is the analysis of individual cells as they pass through a fluid stream. Components of the cells are labeled with one or more fluorescent dyes to identify the cells. A laser within the flow cytometer excites the fluorescent molecules to produce light at numerous wavelengths. The amount of fluorescent light that is emitted from the study is an indication of the type of cells that are within the sample. Flow cytometry is often used during cancer analysis, immunology and cell cycle analysis.

One of the primary steps required to perform cytometry analysis is the staining of single cells with fluorescent dyes. Labeling on the cells is based on whether the cells have existing fluorescent dye. Florescent dye labeling will be based on the availability of antibodies within the proteins. If the antibodies exist than the cells will be stained using antibodies. If the cells do not contain antibodies, than the cells will be stained with an unlabeled antibody, then a second staining will occur with the florescent labeled antibodies.

After the cells have been classified, flow cytometry analysis can be performed. The suspension cells will be loaded into a tube and placed into the flow cytometer. The cells will flow through the instrument and pass in front of the laser individually. As the cell is hit by the laser, light will scatter mainly in two directions. The light that scatters toward the front of the cell will indicate the cell size, while the side scatter will measure the quality of the elements within the cell.

The main purpose of the laser is to excite the florescent molecules within the cells. The light emitted from the processes is at a 525 nm. The photomultiplier tubes within the cytometer contain light filters that are able to identify the light produced by the florescent dye. The cytometer may have several photomultiplier tubes to identify the various wavelengths produced by the fluorescent molecules.

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