Bio-Rad iQ™5 Optical System Software, Version 2.1 - Manual (Page 85)

Section 6 Data Analysis Module
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Positives & Negatives: Select this method to use positive and negative controls to
define/call unknown samples. Samples are considered positive if they are greater in RFU
value than the positive control average minus the tolerance. Samples are considered
negative if their RFU value is less than the negative control average plus the tolerance
End Point Tolerance
End Point Tolerance defines the margins for sorting unknowns as positives or negatives. How the
tolerance variable and the type of tolerance are applied depends on which method is selected.
The End Point Tolerance drop-down list box consists of two choices:
• RFUs: This is the default Tolerance choice and should be selected if you would like to
use an absolute RFU value for the tolerance value. The minimum RFU tolerance value is
2, whereas the maximum is the absolute value of the highest RFU value minus the
absolute value of the lowest RFU value. The default RFU tolerance value is 10% of the
total RFU range. The definition of the RFU range is dependent on the method chosen
•
For the negative method, the range is the highest RFU value minus the negative control
average.
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For the positive method, the range is the positive control average minus the lowest RFU
value.
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For the positives & negatives method, the range is the positive control average minus the
Negative control average.
•
Percent of Range: Select this setting if you would like to use a percentage of the RFU
range for the tolerance value. The minimum percent of range is 1 percent, whereas the
maximum percent of range is 99 percent. The default percent of range tolerance is 10
percent.
End Cycles to Average
End Cycles to Average is the number of cycles from the last cycle, which will be used to calculate
an average End Point RFU value. The End Cycles to Average field defaults to 2 for End Point Only
runs and 5 for non-End Point Only runs. In order for the iQ5 software to analyze non-End Point
Only data in the End Point tab, at least 6 repeats of data acquisition must be performed.
Number of Ranks
The Number of Ranks allows assignment of samples into distinct groups based on their RFU
values. The absolute range (highest RFU minus lowest RFU) is divided by the number of ranks
selected. The default rank value is 10 and the minimum number of ranks is 3.
Colored rank boxes, displayed below Number of Ranks and Sort Data by Call, symbolize the
number and order of ranks in the end point analysis. To the right of the colored rank boxes are
five color-gradation buttons that allow a change in the color scheme of the rank boxes, once the
data are analyzed.
Sort Data by Call
This function is used to sort End Point samples into Positive and Negative Calls (as well as No
Calls wells which do not fall into the previous two categories). Sort Data by Call is essentially the
Positives & Negatives method but also includes sorting and color coding the Positive and
Negative Calls. The ranking function is disabled in this mode.