Fluid Imaging Technologies FlowCAM - Bead Size and Count Interpretation

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Filter Name
Min. ESD Diameter
(µm)
Max. ESD Diameter
(µm)
5µm 3 7
10µm 8 12
20µm 17 23
50µm 46 54
100µm 90 110
300µm 280 320
Bead Size Procedure: The acceptable region (Setup > Context > Capture > Acceptable Region) should
be constrained by one pixel on each side. If a field-of-view (FOV) flow cell is being tested, then adjust
those values accordingly. Dispense 0.5 mL of distilled water into the unit, and prime the system until
the water is past the bottom of the flow cell. Ensure no bubbles are stuck in the flow cell. Set the pump
speed to the appropriate settings for the objective and flow cell under test. Dispense 1 mL of sample for
20x, 10x, and 4x objectives or 3 mL for 2x objectives into the FlowCAM. Start an AutoImage analysis
and save the file with an appropriate name (i.g. Instrument Serial Number_Date_Size_Run Number). If
necessary start the pump and run the sample through until the trailing meniscus reaches the top of the
flow cell. Stop the run before the meniscus enters into the camera’s field of view, to prevent the
erroneous imaging of bubbles.
Interpretation of Results: Each of the six filters (when selected) will display the instrument’s ability to
measure the beads captured. On the Main Window, click each appropriate bead size and note the peaks
that are highlighted on the Frequency vs. Diameter graph. If the highest peak at each measurement is
highlighted, then the FlowCAM/VisualSpreadsheet measured the beads correctly. See the example
below:
The above graphics indicate that the instrument has correctly measured 50µm beads. There will be
slight variation in bead measurements around each size grouping due to the quality of focus and location
within the viewing window that the bead was imaged. This is a normal and acceptable characteristic.
Repeat this data review process for each bead size in the sample. Use the Test Results Checklist at the
end of this Appendix to record the results and store as part of the instrument history record. If the
instrument does not measure the beads correctly (within the error values stated by the microsphere