Troubleshooting
Observation
Possible cause
Recommended action
Uneven screen illumination
(screen is dark on one side,
bright on the other side)
Light cubes misaligned.
Reset the light cube tray by selecting Change
light cube from the Instrument Settings screen.
See Appendix C, “EVOS light cubes”.
Autofocus does not focus
properly
Debris or other material
interfered with the focus.
Make sure there are no bubbles or debris visible
on the screen that could interfere with the
autofocus and make it more difficult to get the
sample in the correct focal plane.
Live cells vs dead cells were
difficult to distinguish.
If cells are well focused, have bright centers, and
are being counted as dead, confirm that they
are within the appropriate cell size range and try
adjusting the settings.
Some cells appear in the image
but are not included in the count
Brightfield or fluorescence
settings were not optimal.
For cell count and cell viability assays performed
in brightfield, adjust the size, brightness, and
circularity gates for both live and dead cells to
include all of the cells in the count.
For fluorescence assays, adjust the size,
brightness, circularity, and fluorescence intensity
gates in all available channels to include all of
the cells in the count.
After including all of the cells in the count,
you can narrow the count criteria, if you wish
to exclude cells of a certain size or certain
brightness.
When the gates are fully maximized, the CSV
should indicate 0–70 for cell size and 0–255 for
brightness.
Be sure channels are optimally illuminated
in both brightfield and fluorescence (only in
Countess
™
3 FL Automated Cell Counter)
modes.
Fluorescence is extremely bright
and bleeding through into other
filters
Light intensity was not set
correctly.
Decrease the fluorescence light intensity before
counting the cells.
A
56
Countess
™
3 FL Automated Cell Counter User Guide
Summary of Contents for AMQAF2000
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