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Figure 3.2 Laser Beam Shaping
b
c
d
1. Violet laser beams
2. Blue laser beam
3. Red laser beam
Cell Illumination
As cells in the sample stream go through the sensing area of the flow cell, the elliptical laser beam
illuminates them. The cells scatter the laser light and emit fluorescent light from fluorescent dyes
attached to them.
Forward Scatter
The amount of laser light scattered at narrow angles to the axis of the laser beam is called forward
scatter (FS). The amount of FS is proportional to the size of the cell that scattered the laser light.
Side Scatter and Fluorescent Light
The amount of laser light scattered at about a 90° angle to the axis of the laser beam is called side
scatter (SS). The amount of SS is proportional to the granularity of the cell that scattered the laser
light. For example, SS is used to differentiate between lymphocytes, monocytes, and granulocytes.
In addition to the SS, the cells emit fluorescent light (FL) at all angles to the axis of the laser beam.
The amount of FL enables the instrument to measure characteristics of the cells emitting that light,
depending on the reagents used. For example, FL is used to identify molecules, such as cell surface
antigens.
773232AH
e
f
4. Horizontal beam shaping lens
5. Vertical beam shaping lens
Operation Principles
3
Cell Illumination
3-3
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