The Range of the EZ-Fluo™ method ca

The Range of the EZ-Fluo™ method can be compared to the Range of the traditional method.
If it is not possible to determine the Range of the EZ-Fluo™ method, verify that the conditions of Accuracy, Precision and Linearity were met.
EZ-Fluo™ Method Specificity
The EZ-Fluo™ method is specific to viable micro-organisms such as bacteria, yeasts and molds.
The Specificity of EZ-Fluo™ method can be established from the data recovered during the Incubation Time Robustness and Accuracy tests where several micro-organism species were tested.
To demonstrate the EZ-Fluo™ method’s Specificity it is possible to screen the method with the appropriate test micro-organisms.
Several replicates should be carried out for each microorganism.
These tests conditions are covered by the Incubation Time Robustness test, as well as the Accuracy test, where several micro-organisms were challenged.
In these tests the EZ-Fluo™ method is tested in parallel with the traditional method, and microbial recovery is calculated for each micro-organism tested.
Incubation Time Robustness demonstrates that the micro-organisms tested are successfully detected and recovery of the micro-organisms is assessed.
Accuracy test demonstrates that the microorganisms tested are successfully detected and recovery percentage shows closeness of the EZ- Fluo™ results to the traditional Method results.
It is possible to use Appendix 13:Comment Sheet, to document the list of the micro-organisms tested and successfully detected with the EZ-Fluo™ method.
The Specificity of the EZ-Fluo™ method is assessed if all test micro-organisms are successfully detected, where conditions of Robustness, Accuracy, Precision and Linearity are met.
Remark:the specificity can also be assessed from the test 6.10:Verification of the Product Absence of Interference, were different micro-organisms are tested in presence of test products.
EZ-Fluo™ Method False Positive Rate
A False Positive normally means that a test claims a sample to be positive, when it actually is not, and is evaluated where no positives are present in the samples.
An approach to evaluate the occurrence of False Positive is to consider all the negative controls(samples without micro-organism)processed during Incubation Time Robustness and Accuracy tests.
This approach enables comparing the EZ- Fluo™ method and the traditional method, as both are tested in parallel.
The Negative Controls are processed during handlings to verify that the operating conditions are aseptic, and that the method does not generate false positive result.
In case of fluorescent spot on an EZ-Fluo™ method Negative Control sample, an investigation should be carried out.
The investigation plan below is proposed:
EZ-Fluo Negative Control
Fluorescent spot
INVESTIGATION:
Re-incubation of the sample
=Real Positive result due to exogenous contamination
Note:The EZ-Fluo™ sample should be re-incubated to achieve the incubation period recommended by the relevant pharmacopeia.
The False Positive rate of the traditional method should have been determined during the traditional method validation.
If not, it is possible to use the results obtained during the present EZ-Fluo™ method validation, as both methods are tested in parallel.
If using the sampling plan proposed for Incubation Time Robustness and Accuracy test, the following numbers of Negative Controls without micro-organism are performed during each handling set with micro-organisms:
Number of Negative Controls(Per micro-organism tested)
EZ-Fluo™ Method
Incubation Time Robustness Test
Total Number of Negative Controls
In both Incubation Time Robustness and Accuracy tests, the sampling plan is proposed as an example, to test one micro-organism type.
So the more micro-organisms are tested, the more Negative Controls are processed.
These Negative Controls results should be summed, in order to determine the occurrence of false positive result.
The False Positive Rate of the EZ-Fluo™ method can be calculated as follows:
Number of False Positives Spots
The False Positive Rate of the EZ-Fluo™ method is compared to the False Positive Rate of the traditional method.