Bio-Rad VINEO™ Brettanomytest PCR Kit - Manual (Page 9)

You must use the the Opticon Monitor™ software (version 3.3 and higher)
or the CFX Manager™ software Industrial Diagnostic Edition for an
automated and quantitative interpretation. A complete report may be printed
(cf. software instructions).
1. Controls
Before a final interpretation of the results, you have to check the results of the
negative and positive controls.
For the test to be valid, the results of the negative and positive controls must
be as follows:
* N/A means “Not Applicable”. The software indicates N/A for the Ct of a sample when the
fluorescence curve does not cross the threshold.
If the results of the positive and negative PCR Qs controls are different from
those described in the table below, you have to begin the PCR again.
2. Samples
A sample is considered to be positive for Brettanomyces bruxellensis if the
value of Ct ≥ 10 is obtained for the FAM fluorophore.
If no value is obtained for Ct
FAM
(Ct
FAM
= N/A), the interpretation of the result
depends on the value of the internal control:
- A sample is considered to be negative for Brettanomyces bruxellensis if
no value of Ct is obtained for the FAM fluorophore (Ct
FAM
= N/A and if the
Ct of the internal control is greater than or equal to 28. (Ct
HEX
≥ 28).
- An N/A value for the Ct
HEX
of the internal control indicates that an
inhibition phenomenon of the PCR reaction has probably occurred, if
the Ct
FAM
of the target is also N/A. In this case, the DNA sample must
be diluted (to 1/10 th for example) in sterile distilled water and then
subjected to a new PCR.
- If the Ct
HEX
of the internal control is less than 28, it is impossible to
interpret the result. Check that the threshold was set correctly or that
the raw curve shows an aspect characteristic of exponential
amplification. If the observed curve is not correct, it will be necessary to
repeat the PCR test for this sample.
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Detection of Brettanomyces
bruxellensis (FAM)
Detection of the internal
control (Channel HEX)
Negative control Ct = N/A* 25 ≤ Ct ≤ 40
Positive Qs PCR control 27 ≤ Ct ≤ 37 Not significant