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The specificity of detection is controlled by optical filters. The detectors in most flow cytometers are usually PMTs. By combining the FSC and SSC information with fluorescence labeling, it is possible to differentiate cell types in a heterogeneous population such as blood. FSC data provides an estimate of a particle's size while SSC provides information on the relative internal complexity of a cell. The optical output is collected as forward or side scattered light (FSC or SSC) by a PMT or photodiode and the optical data is used to characterize the cell properties. The third system is the electronics and signal processing equipment that converts the detected optical signals to electronic signals for processing and analysis.įollowing hydrodynamic focusing, the particle stream passes through one or more focused laser beams and light scattering or fluorescence emission occurs. The second is the optical system composed of lasers or light sources that illuminate the particles in the sample stream as well as optical filters and beamsplitters that direct the post-sample light signals to optical detectors for counting and processing. The first is the fluidics system that transports sample particles to the laser beam in a narrow, single particle wide stream.
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All photonics components must be precisely aligned and controlled with µm spatial resolution.Ī flow cytometer (Figure 1) consists of the three major component systems. Dichroic filters with precision coatings are needed for narrow reflection/transmission bands and high optical density for out-of-band rejection. Laser light with specific wavelengths is needed to excite labels or fluorophores on a cell. The method requires several photonics components, starting with lasers and other light sources with small, tightly focused beams that can illuminate a single size cell or particle (~30 µm diameter). The properties measured include relative particle size, relative granularity or internal complexity, and relative fluorescence intensity. A flow cytometer takes a sample of cells, transitions them into a single stream and uses lasers and/or light sources to excite biomarkers or labels on the cells to count the number of relevant constituents. Flow cytometry is an analytical technique that can rapidly measure the properties of individual cells or particles as they pass through a beam of light, typically a laser.