Laser Beam Shaping and Spot Size

The majority of fluorescence flow cytome-ters now in use derive illumination from one or more lasers. Air-cooled argon ion lasers operating at 488 nm are the most commonly used light sources, but helium-neon (He-Ne) lasers emitting at 633 nm and diode lasers emitting at 635 nm have come into more widespread use as red-excited antibody labels and other fluorescent probes have become available. Other lasers sources used in fluorescence flow cy-tometry include argon, krypton, and mixed-gas ion lasers operating at wavelengths from the ultraviolet to the infrared; helium-cadmium (He-Cd) lasers emitting in the ultraviolet region at 325 nm and/or in the blue-violet region at 441 nm; and diode-pumped, frequency-doubled yttrium aluminum garnet (YAG) lasers emitting green light at 532 nm. A typical laser emits a beam on the order of 1 mm in diameter; efficient use of the laser light requires that this beam be shaped and focused to a smaller size to illuminate the cell stream passing through the flow cytometer. This unit will discuss spot size requirements and the methods by which optimal spot size is achieved.

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