129
Overview of procedure:
The experiment is set up using the Quantification Wizard as detailed above. The experiment must
include a dilution series of standards in the plate layout by which to compare the unknowns. The
concentration of each standard should be added to the
Well Information
table in the Plate Layout
Editor (can be accessed through the Results Editor main screen, post-run).
•
If less than two standards have been assigned in the plate layout, a message in the Results
Editor will alert the user to the fact that there are insufficient standards.
•
If no standards have been assigned, no standard curve graph will be displayed.
4.6.6.1
Quantification cycle graph (fit points analysis only)
•
Click the Cq icon if the quantification cycle graph is not displayed.
The results (with or without correction) are plotted on a scale of log fluorescence vs. cycle number.
Although some fluorescence values may be negative, this has no effect on the accuracy of the
data.
The blue line represents the
noise threshold and the red line
the crossing line.
The user can manually adjust
the position of the lines by
dragging with the mouse to
positions better suiting the data.
The position can also be
changed from the PAR box
accessed by clicking on the
PAR
button next to the Cq
graph.
Changing the position of the cursors will update the graphs. This can be useful for adjusting the
best fit such that the R
2
value (correlation coefficient) is as close to 1 as possible i.e. a perfect
correlation (see The standard curve below).
Cycle Number
40
35
30
25
20
15
10
5
R
e
la
ti
v
e
F
lu
o
re
s
c
e
n
c
e
(
L
o
g
)
0.0001
0.001
0.01
0.1
1
10
