ChemoMetec NC-100 - Manual (Page 25)

2 Introducing the NucleoCounter
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by multiplying the imaged area with the depth of the cassette. The imaged area is only
dependent on the optics of the instrument. Therefore, the area is constant and specific
for each instrument.
2.2.4 Cassette handling
The built-in image analysis method of the NucleoCounter offers considerable stability,
when eliminating non-cellular objects from the image, including scratches and smears
on the cassette windows, and thus producing valid results even under extreme
conditions. On the other hand, the quality of the cell count is best assured by keeping
the windows of the cassette as clean as possible.
Caution! In order to avoid contaminating the measurement window it is important not
to touch the window when handling the cassettes. Be careful when attempting to wipe
off the surface of the cassette, in an attempt to remove any foreign object, since the
plastic material of the cassette can be scratched.
2.3 Lysis buffer (Reagent A100) and Stabilizing buffer (Reagent B)
The Lysis buffer is used for disruption of the plasma membranes, rendering the nuclei
susceptible to staining with propidium iodide. Furthermore, when using the Lysis buffer,
the cell aggregates are dissolved.
The main objective of the Stabilizing buffer is to raise pH of the sample mixture,
thereby optimizing the fluorescence of propidium iodide and to stabilize the cell nuclei.
The combination of treating the cell sample first with the Lysis buffer and then
Stabilizing buffer is ideal for the total cell count, since staining of the nuclei is optimal
under these conditions.