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A TELIERS INTERACTIFS
customer base then this particular risk element may be
lower.
When it comes to actually implementing a Rapid
Microbiology Test System a number of criteria will
become very important and the ability of the supplier
to provide these should be established at the audit
stage.
Procedures and documentation for the following should
be available
1. Installation 4. Calibration
2. Operation 5. Service
3. Training
So finally the responsibility of the equipment supplier is
to validate their system according to USP or
, or PDA Technical Report 33 and be able to
provide the data to demonstrate this and to facilitate
customers to do the same.
Using the Validation Guidelines
USP /PDA TR33: Validation of Alternative
Microbiological Methods
These chapters give clear guidance on the following criteria
1. Accuracy 5. Linearity
2. Precision 6. Range
3. Specificity 7. Ruggedness
4. Quantification Limit 8. Robustness
I will discuss some of these criteria using the example
of the Millipore Microstar System. The Microstar is a
complete system for the detection and enumeration of
viable microorganisms in filterable samples. Typically
it takes only 1/4 to 1/3 the time of conventional
methods to achieve a comparable result.
The Microstar uses a traditional microbiology technique
(membrane filtration) coupled with ATP bioluminescence
and CCD image analysis to give you comparable
results to the compendial method, but within a
much shorter time frame. The Microstar can be used
for testing water, in-process product and final product
within the Pharmaceutical Industry.
Accuracy
Accuracy can be defined as the closeness of a test
result to the results predicted by the compendial
method. Accuracy is expressed as the percentage recovery
of microorganisms by the test method.
The alternate method must provide an estimate of not
less than 70% of the estimate provided by the compendial
method.
An example is shown below:
Organism Average Percent Recovery
E. coli 95
A. niger 127
P. aeruginosa 82
C. albicans 104
S. aureus 97
B. subtilis 108
Precision
Precision can be defined as the degree of agreement
among test results when the procedure is applied to
multiple samplings of suspensions of microorganisms
across the range of a test. These have been shown to be
equivalent when using Microstar to compendial
methods
Specificity
Specificity can be defined as the ability to detect a
range of microorganisms, which demonstrate that the
method is fit for purpose.
The method’s compatibility with the different types of
sample matrices should also be shown.
To determine this screen the method against a representative
range of microorganisms and a range of sample
types that are appropriate to the method.
Acceptance Criteria: All microorganisms selected as
representative are successfully detected and enumerated
in the sample matrices. The Microstar System has
successfully been used to detect a very wide range of
organisms in a huge range of sample matrices ranging
from the straightforward such as water to the more
challenging such as mammalian cell culture suspensions
Quantification Limit
Quantification Limit can be defined as the lowest number
of microorganisms that can be determined with
acceptable precision and accuracy under the stated
experimental conditions.
Z-Statistic and p-value determine whether or not data
are statistically similar. The following table illustrates
this.
Organism MicroStar HA MF Statistically
Similar?
E. coli 10.6 9.7 Yes
A. niger 9.6 8.2 Yes
P. aeruginosa 8.8 9.1 Yes
C. albicans 10.4 8.8 Yes
S. aureus 14.5 12.8 Yes
B. subtilis 8.0 7.0 Yes
Linearity and Range
Linearity can be defined as the ability of the test
method to elicit results, which are proportional to the
concentration of microorganisms present in the sample
within a given range. Microstar shows excellent linearity
up to counts of around 100 cfu per membrane. If
higher counts than this are expected a dilution step
may be recommended.
Ruggedness
Ruggedness can be defined as the degree of precision
of test results obtained by analysis of the same samples
under a variety of normal test conditions, such as different
analysts, different instruments, different lots of
reagent, etc. It is often best suited to determination by
the test supplier who has easy access to multiple instruments
and batches of components. Millipore has
conducted extensive testing using different reagent lots
on different Microstar equipment. Extensive shipping
trials have also been conducted on both reagents and
equipment
Robustness
Robustness can be defined as a measure of a method’s
capacity to remain unaffected by small, but deliberate
variations in method parameters, and provides an indication
of its reliability during normal usage. Again this
is often best suited to determination by the equipment
supplier. Millipore has validated many areas of robustness
in relation to the Microstar including false positive
rates and incubation times. The Microstar method has
been shown to be highly robust.
Finally to summarise, the implementation of a Rapid
Microbilogical Test Method can be achieved much more
smoothly by first of all a careful audit of the suppliers
capabilities and then by using these capabilities in
conjunction with the available regulatory guidelines to
validate your new test method. ■
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