ChemoMetec NC-100 - Manual (Page 42)

6 Total and viability count
26
6.3 Calculate viability
When both the concentration of non-viable cells and the total concentration of cells are
known, it is possible to calculate the viability (%viability) of the cells. The calculation
can either be done manually or by use of the NucleoView software (please refer to the
NucleoView
TM User´s Guide). For the calculation of %viability the multiplication factors
must be taken into account according to the equation shown below.
100%
M C
M C M C
%viability
t t
nv nv t t
⋅
⋅
⋅ − ⋅
=
%viability: The percentage of viable cells in the original cell suspension.
C
t
: The total concentration of cells in the NucleoCassette (the displayed
result of the total cell count).
C
nv
: The concentration of non-viable cells in the NucleoCassette (the result
displayed when counting the non-viable cells).
M
t
: The multiplication factor used for the total cell count.
M
nv
: The multiplication factor used for the counting of non-viable cells (most
often 1).
Example of calculation of %viability: Example of calculation of %viability: Example of calculation of %viability: Example of calculation of %viability:
The stabilized lyzate is prepared using 200 µl of the cell suspension, 200 µl Lysis buffer
and 200 µl Stabilizing buffer. Thus, the multiplication factor (M
t
) is 3. The result of the
total cell count (C
t
) is 6,7•10 5 cells/ml.
Counting the non-viable cells is done without dilution of the cell suspension prior to the
analysis and the multiplication factor (M
nv
) is 1. The result of the count of non-viable
cells (C
n v
) is 3,0•10 4 cells/ml.
The calculation of %viability is shown below.
98,5% 100%
3 10 6,7
1 10 3,0 - 3 10 6,7
100%
M C
M C M C
%viability
5
4 5
t t
nv nv t t
= ⋅
⋅ ⋅
⋅ ⋅ ⋅ ⋅
= ⋅
⋅
⋅ − ⋅
=
6.4 Dilute the samples
As mentioned earlier the NucleoCounter is able to count cells in the range 5x10 3 - 2x10 6
cells/ml, the optimal range being 1x10 5 – 2x10 6 cells/ml. Therefore it can be necessary
to dilute or concentrate the cell samples. There are two recommended methods to
dilute a cell sample; either it can be diluted using the Lysis buffer and the Stabilizing
buffer or it can be diluted prior to addition of Lysis buffer and Stabilizing buffer.
When using the Lysis buffer and Stabilizing buffer to dilute the cell sample, the
procedure described above is followed, only the amounts of Lysis buffer and Stabilizing