ChemoMetec NC-100 - Manual (Page 43)

6 Total and viability count
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buffer added to the samples are altered. Note that it is important to add equal volumes
of Lysis buffer and Stabilizing buffer to the samples. For examples of dilution using the
Lysis buffer and Stabilizing buffer, see the table below.
Cel Cel Cel Cel l sample l sample l sample l sample Lysis buffer Lysis buffer Lysis buffer Lysis buffer Stabilizing buffer Stabilizing buffer Stabilizing buffer Stabilizing buffer
Multiplication Multiplication Multiplication Multiplication
factor factor factor factor
200 µl 200 µl 200 µl 3
200 µl 400 µl 400 µl 5
200 µl 600 µl 600 µl 7
200 µl 800 µl 800 µl 9
When it is desired to dilute the stabilized lyzate it is recommended to mix equal
portions of Lysis buffer, Stabilizing buffer and cell culture media (optionally a balanced
salt solution such as PBS) and then dilute the stabilized lyzate with the mixture.
It is also possible to dilute the cell sample prior to treatment with Lysis buffer and
Stabilizing buffer using either cell culture media or a balanced salt solution. When using
this method to dilute a cell suspension, take out a sample of the diluted cell suspension
and add Lysis buffer and Stabilizing buffer in the normal way. Finally calculate the
multiplication factor.
Example of multiplication factor calculation: Example of multiplication factor calculation: Example of multiplication factor calculation: Example of multiplication factor calculation:
The cell suspension is diluted 5 times by taking out 200 µl and adding 800 µl PBS. Then
a 200 µl sample is drawn from the diluted suspension, 200 µl Lysis buffer and 200 µl
Stabilizing buffer is added. Thereby the sample is diluted another 3 times. The cells are
counted on the NucleoCounter.
The multiplication factor is calculated from the two separate dilutions. In this example
the multiplication factor is 5x3 = 15. Therefore the concentration of cells obtained from
the NucleoCounter has to be multiplied by 15 to calculate the concentration of cells in
the cell suspension.