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Sanas TR 26-02
Sanas TR 26-02
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TR 26-02
CONTENTS
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The purpose of this document is to define the concepts and processes of method validation, and to
provide technical requirements in order to facilitate a uniform approach. This document amplifies
ISO/IEC 17025:2005 requirements and lists SANAS requirements applicable to chemical laboratories.
The following do not form part of this document: Verification, sampling, sample handling and
transportation.
2.1 References
a) Eurachem Guide. The Fitness for Purpose of Analytical Methods. A laboratory guide to
method validation and related topics. Copyright LGC (Teddington) Ltd 1998
b) ISO/IEC 17025. General Requirements for the competence of testing and calibration
laboratories.
c) Ludwig Huber, Validation and Qualification in Analytical Laboratories, second edition. Ludwig
Huber. Agilent Technologies. Waldbronn, Germany.
d) TG 41-01 “Recommended guidelines for the verification and validation of methods in forensic
chemistry”.
e) Quantitative Chemical Analysis Second Edition Gilbert H. Ayres, Prof of Chemistry,
University of Texas at Austin
f) PALCAN Guidance for the Validation of Test Methods. Can-P-1629. November 2009.
Standards Council of Canada.
th
g) Skoog et al, Fundamentals of Analytical Chemistry, 7 Edition, 1996.
2.2 Definitions
2.1.1 Accuracy - The accuracy of an analytical method is the extent to which test results
generated by the method and the true value agree. Accuracy can also be described as
the closeness of agreement between the value that is adopted, either as a conventional,
true, or accepted reference value, and the value found. (3)
2.1.2 Precision - The closeness of agreement between independent test results obtained
under stipulated conditions (how close the measured values are to each other). It is
usually expressed as the standard deviation or relative standard deviation (co-efficient of
variance) and may be a measure of either the degree of reproducibility and /or
repeatability (1).
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The required precision for a method is determined by the role the test results are
going to play in making the decision regarding the release of the product.
Precision may not be relevant to an identification test, but if may be highly relevant
to an assay.
It must be noted that the RSD is usually high at low concentrations and low at
higher concentrations.
ii) The analysis is carried out in one laboratory by one operator, using
one piece of equipment, over a relatively short time span (normally
one day).
i) A regression line has errors in the slope and the intercept and when
an unknown concentration (x0) is determined by using the regression
line, a regression error (Sx0) is present.
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ii) A typical criterium for regression precision would be RSD < 2.5%.
2.2.2.4 Ruggedness
ii) A rugged method is one that has built in buffers against typical
abuses, that is, against differences in care, technique, equipment,
and conditions.
ii) If a true (accepted) value is not available apply the paired t test and
F test to the results generated by different analysts.
iii) If the calculated F value exceeds the tabulated F value at the selected
confidence level, then there is a significant difference between the
variances of the two analysts.
iv) The statistical calculated paired t value shall not exceed the tabulated
value at the desired confidence level.
2.2.2.6 Reproducibility
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2.2.3 Bias – It is the tendency of a method towards delivering a result that is skewed from the
true value. It is the difference between the experimental mean and the true value and is
generated from a total systematic error as contrasted to random error. There may be one
or more systematic error components contributing to the bias.
2.2.4 Reproducibility - This refers to replicate analysis performed using the same method on
identical test items using different operators and/or instruments and/or laboratories over a
longer interval of time
2.2.5 Repeatability - This refers to replicate analysis performed using the same method on
identical test items using the same operator, the same instruments and within the same
laboratory over a short interval of time.
2.2.6 Linearity - Ability of a method to obtain test results proportional to the concentration of
the analyte within a given working range.
2.2.7 Working range - The range of an analytical method is the interval between the upper and
lower levels of an analyte, including these levels that have been demonstrated to be
determined with a suitable level of precision, accuracy and linearity, using the method as
written. The working range is normally expressed in the same units as the test results
obtained by the analytical method.
2.2.8 Limit of Detection (LOD) - The lowest concentration of analyte that can be detected but
not necessarily quantitated under the stated conditions of the test. It is a point at which a
measured value is larger than the uncertainty associated with it.
2.2.9 Verification - Confirmation by examination and provision of objective evidence that
specified requirements have been fulfilled 1).
2.2.10 Uncertainty of Measurement (Measurement Uncertainty) - Parameter associated with
the result of a measurement, that characterizes the dispersion of the values that could
reasonably be attributed to the measurand (1).
2.2.11 Limit of Quantification LOQ - The lowest concentration of analyte that can be
determined with acceptable precision (and accuracy) under the stated conditions of the
test (4).
2.2.12 Sensitivity - Capability of the method to discriminate between small differences of
concentrations of analyte.
2.2.13 Specificity / Selectivity - The ability of a method to respond to a particular analyte of
interest in the presence of possible interferences such as impurities, degradents and
matrix effects.
Figure 2: The Definitions for linearity, range, LOQ and LOD displayed.
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Figure 3: The definitions for LOD and LOQ displayed: Limit of detection and limit of
quantitation via signal to noise.
3. Introduction
3.1 Method validation is the process of establishing the performance characteristics and limitations of
a method and the identification of the influences which may change these characteristics and to
what extent. It is also the process of verifying that a method is fit for purpose, i.e. for use for
solving a particular analytical problem (1).
3.2 Verification refers to a process that provides evidence that the laboratory can achieve the
performance characteristics given in a specific analytical method, especially accuracy and
precision, and demonstrating that the method is suitable for the intended use. The extent and
nature of such verification work depends on the needs of the customer, and the intended use (4).
3.3 Validation is always a balance between costs, risks and technical possibilities.
3.4 For clarification on when to perform validation or verification see Table 1. This Table has been
adapted from PALCAN Guidance for the Validation of Test Methods (3).
Table 1: When should methods be validated? (3)
Standard published method plus Full validation may be required only if changes made.
additional documentation for optional
steps.
Changes in implementation of previously Extent of validation will vary to demonstrate change does
validated methods – i.e. changes to not have a significant impact on performance
equipment, reagents, lab environment or characteristics.
staff.
Standard published method applied to Validation is required and the extent will vary, e.g. having
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different matrices, different concentration similar properties to those of representative matrices and
ranges, analytes or standard published analytes.
method used for a similar purpose but
different conditions.
4.1 Performance characteristic “means functional quality that can be attributed to an analytical
method” (EC Directive). Examples of typical performance characteristics include: selectivity,
accuracy, trueness, recovery, precision, repeatability, reproducibility, detection limit, limit of
quantitation, detection capability, ruggedness and stability. The laboratory may also be required
to evaluate sampling, subsampling and transportation of samples to the laboratory as part of the
validation plan.
4.2 Performance criteria “means requirements for a performance characteristic according to which it
can be judged that the analytical method is fit for the purpose and generates reliable results.” (EC
Directive).
5. Validation Plan
5.1 The scope of the method and its validation criteria must be defined early in the process. These
include the following questions:
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Note: How test method performance characteristics are evaluated, along with the criteria against which
they will be assessed, are usually described in your standard published methods, scientific literature or
equipment specifications. It is therefore the responsibility of the laboratory, with input from clients, to
seek out the relevant characteristics to be evaluated with respect to the laboratory’s specific situation
and client’s needs. The laboratory must have a documented validation plan, either to be used generally
or applied to a specific project or client. Test method performance characteristics to be evaluated will
vary with the type of test and its intended use. Discipline-specific or client required performance criteria
are to be applied to demonstrate fitness for purpose. (3)
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6.1 Analyse the data applying the appropriate statistical tools, e.g. Analysis of Variance (ANOVA tool,
on Excel, for statistics), linear regression, t- test, f- test, etc.
Note 1: Ensure that conventions regarding use of significant figures and rounding data is applied
before analysing the data. Remember that when rounding data from chemical computations it
may be necessary to carry one extra digit through all the computations to avoid rounding error. In
rounding a number ending in 5, always round so that the result ends with an even number. (7)
Note 2: Units give dimensions to numbers therefore do not forget your measurement units.
7. Summary Report
7.1 The laboratory must have available for review a report, summarizing all the detailed method
validation data for all non-standard, in-house developed or modifications and amplifications of
standard published methods. The report should include:
7.2 The test method as validated. This includes information about equipment, reagents, calibration
etc. (Confusion may arise if the method does not meet performance criteria and further method
development is required).
7.3 Reference to the validation procedure or plan used to generate the test method performance
characteristics.
7.4 A summary of the test method performance characteristics and how these were calculated or
defined. The raw data should also be available for review.
7.5 The test method performance criteria against which the characteristics were evaluated and
whether or not the method is fit for purpose.
7.6 The intended use of the method.
7.7 Estimates of uncertainty based on interpretative documents of the ISO GUM (for example, the
EURACHEM CITAC Guide).
7.8 If the method that is not a standard published method is used routinely, it is expected that over
time there will modifications or improvements made. This information needs to be documented
and available for assessment. Ongoing proficiency testing data and quality control data should be
reviewed by the laboratory to confirm the fitness of the method.
7.9 SANAS requires is that all validation data should be readily available in the laboratory for a
minimum of at least one assessment cycle (5 year cycle).
8. Revalidation
The laboratory is expected to continually prove that the validation is still current through the quality control
procedures that include the method’s performance characteristics/parameters. Part or full re-validation
may be considered when:
ii) new samples with new compounds or new matrices are introduced (Huber: 8.3),
iii) a new location with different environmental conditions is used (Huber: 8.3),
iv) new chemicals and/or reference standards are used (Huber: 8.3),
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vi) a review of quality control indicates an established method is changing with time,
viii) in the case of the method performance criteria falling outside the acceptance criteria (Hube 8.3)
Note : In the case where a new analyst is appointed to perform analysis, the laboratory is expected to
ensure that the analyst is competent and meets the method’s/relevant procedure’s performance criteria.
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