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Advanced Overview of the FlowCAM
The FlowCAM architecture can be divided into three distinct systems:
· Optics
· Fluidics
· Electronics
Optics
Figure 1 is a block diagram of the FlowCAM architecture. The following
section explains the operation of the components illustrated when using laser
dependent Fluorescence and/or Scatter Trigger modes.
As the particles are drawn into the flow chamber they pass through a “laser
fan” that has been generated by the laser and a lens. Fluorescence which
results from this illumination is passed back through the objective and the
partial mirror. This light then passes through a system of optical filters and
mirrors. The shorter wavelengths of green to orange are reflected by a mirror
to the 500-600 nm fluorescence detector. Longer wavelength light passes
through another filter to the 600-700 nm fluorescence detector. Finally, a scatter detector is located at
the end of the optical path (in the same cube as the flash LED) to detect any disturbances in the laser
light.
If either the fluorescence or scatter detector receives a strong enough signal, indicating the presence of a
particle of interest, the flash LED is turned on for a very short interval. This light is then imaged with
the objective onto the camera. A patented depth of focus enhancer stretches out the focus of the
objective, thus enhancing the resolution of particle images.
Fluidics
Sample fluid is drawn through the Flow chamber for analysis by a syringe
pump and then deposited into an outflow collection. FlowCAM’s unique
flow chamber is a key component to this system. Unlike conventional flow
cytometers, the FlowCAM does not use a sheath fluid for hydrodynamic
focusing (laminar flow). Instead, it uses a flow chamber with a variable size
cross section. A complete list of the standard flow cell dimensions
recommended for each objective is available in Appendix I.
The appropriate flow chamber size is chosen by considering the following
interdependent factors:
· Objective Magnification
· Size and Size Range of Particles Under Study
· Desired Flow Rate
· Viscosity of the Fluid
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