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9.2 Viewing Fluorescent Images in the IncuCyte™ FLR
Fluorescence scanning generates both an HD (phase contrast) and fluorescence
image at each scan pattern location. Open the Vessel View window and select
the Image tab. The Image tab in the FLR Vessel View Window provides the User
with a selection of viewing options. It is divided into two Channels: A and B. Use
the corresponding pull-down menus to make your selections. See Figure 9-1.
You can select either Phase Contrast or Fluorescence for each channel. In
Figure 9-1, Channel A is set to Fluorescence, and Channel B is set to Phase
Contrast. The two channels are independent, and the settings for each channel
can be independently adjusted. However, the images from the two channels can
be overlayed using the A/B Slider Bar to form a single, composite image. See
The A/B Slider, page 204.
9.2.1 Fluorescence Settings
The Fluorescence settings allow the User to adjust the DISPLAY characteristics
of fluorescence components within an image. Adjusting the display settings will
affect which components of an image will be displayed as fluorescent, and also
the brightness of those fluorescent images relative to each other. It will NOT
affect the absolute (calibrated) fluorescence values. Calibrated fluorescence
values are measured in terms of pixel brightness using Arbitrary Units (AU‟s).
Figure 9-2 shows the case of Channel A and B both set to Fluorescence. The
“mouse over” Pixel Intensity for fluorescence image NEVER CHANGES. The
Pixel report will change based upon whether or not an image has been
processed.
Figure 9-2. View Pixel Intensity and Coordinates
Selecting a Minimum and Maximum Intensity basically “clips” the brightness at
which fluorescent components will be displayed. In Figure 9-1, the Maximum
Intensity is set to 104.3. Nonetheless, it is possible that the image will still
contain areas with calibrated pixel values greater than 104.3. However, these
areas will be assigned the SAME degree of brightness in the display as an area
with a pixel value equal to 104.3. Thus all areas with a Pixel Intensity of 104.3 or
greater will be displayed as equally bright. To increase the dynamic range of
brightness displayed for areas at 104.3 AU and above, increase the value of the
Maximum Intensity.
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