CEN/TS 16800:2020

SLOVENSKI STANDARD
01-april-2021
Nadomešča:
SIST-TS CEN/TS 16800:2016
Smernica za validacijo fizikalno-kemijskih analiznih metod
Guideline for the validation of physico-chemical analytical methods
Anleitung zur Validierung physikalisch-chemischer Analysenverfahren
Lignes directrices pour la validation des méthodes d'analyse physico-chimiques
Ta slovenski standard je istoveten z: CEN/TS 16800:2020
ICS:
13.060.50 Preiskava vode na kemične Examination of water for
snovi chemical substances
13.080.10 Kemijske značilnosti tal Chemical characteristics of
soils
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.

CEN/TS 16800
TECHNICAL SPECIFICATION
SPÉCIFICATION TECHNIQUE
December 2020
TECHNISCHE SPEZIFIKATION
ICS 13.060.50 Supersedes CEN/TS 16800:2015
English Version
Guideline for the validation of physico-chemical analytical
methods
Lignes directrices pour la validation des méthodes Anleitung zur Validierung physikalisch-chemischer
d'analyse physico-chimiques Analysenverfahren
This Technical Specification (CEN/TS) was approved by CEN on 9 November 2020 for provisional application.

The period of validity of this CEN/TS is limited initially to three years. After two years the members of CEN will be requested to
submit their comments, particularly on the question whether the CEN/TS can be converted into a European Standard.

CEN members are required to announce the existence of this CEN/TS in the same way as for an EN and to make the CEN/TS
available promptly at national level in an appropriate form. It is permissible to keep conflicting national standards in force (in
parallel to the CEN/TS) until the final decision about the possible conversion of the CEN/TS into an EN is reached.

CEN members are the national standards bodies of Austria, Belgium, Bulgaria, Croatia, Cyprus, Czech Republic, Denmark, Estonia,
Finland, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway,
Poland, Portugal, Republic of North Macedonia, Romania, Serbia, Slovakia, Slovenia, Spain, Sweden, Switzerland, Turkey and
United Kingdom.
EUROPEAN COMMITTEE FOR STANDARDIZATION
COMITÉ EUROPÉEN DE NORMALISATION

EUROPÄISCHES KOMITEE FÜR NORMUNG

CEN-CENELEC Management Centre: Rue de la Science 23, B-1040 Brussels
© 2020 CEN All rights of exploitation in any form and by any means reserved Ref. No. CEN/TS 16800:2020 E
worldwide for CEN national Members.

Contents Page
European foreword . 4
Introduction . 5
1 Scope . 6
2 Normative references . 6
3 Terms and definitions . 7
4 Principle . 14
5 Characterization toolbox: performance characteristics . 16
5.1 Introduction . 16
5.2 Characteristics . 16
5.2.1 Selectivity . 16
5.2.2 Sensitivity . 17
5.2.3 Robustness. 17
5.2.4 Trueness . 17
5.2.5 Precision . 19
5.2.6 Limit values . 19
5.2.7 Calibration . 20
5.2.8 Application range . 20
5.2.9 Measurement uncertainty . 20
6 Method development . 21
7 Intra-laboratory validation (V1) – option 1, basic procedure . 23
7.1 General . 23
7.1.1 Validation 1 . 23
7.1.2 Adoption of a standardized method . 23
7.1.3 Extension of the application domain of an intra-laboratory validated method . 23
7.1.4 Complete in-house development . 23
7.2 Intra-laboratory performance characteristics . 24
7.2.1 General . 24
7.2.2 Trueness . 24
7.2.3 Precision . 24
7.2.4 LOD, LOQ . 25
7.2.5 Measurement uncertainty . 26
8 Intra-laboratory validation (V1) – option 2, including verification of the LOQ . 26
8.1 General . 26
8.2 LOQ-V . 26
9 Interlaboratory validation 2 (V2) . 27
9.1 General . 27
9.2 Procedure. 28
9.2.1 Participating laboratories. 28
9.2.2 Materials: selection, preparation and pre-testing of samples . 29
9.2.3 Replicates . 30
9.2.4 Characteristics of the inter-laboratory study . 30
9.2.5 Assigned values . 30
9.2.6 Statistical evaluation and calculation of the results . 31
10 Validation report . 32
10.1 General . 32
10.2 Module A: Test method definition, documentation and general requirements . 32
10.3 Module B: Applicability domain validation . 33
10.4 Module C: Intra-laboratory performance . 33
10.5 Inter-laboratory validation . 33
10.5.1 General . 33
10.5.2 Documentation, publication and standardization . 34
Annex A (normative) Intra-laboratory validation . 35
A.1 Module A: Test method definition, documentation and general requirements . 35
A.2 Module B: Application range and pre-validation . 38
Annex B (normative) Module C: Intra-laboratory performance . 40
Annex C (normative) Module D: Requirements on the study for inter-laboratory validation . 41
Annex D (informative) Structure and content of a validation study documentation (V2) . 44
Annex E (informative) Robustness testing by systematic variation of influencing factors . 49
E.1 Design of experiment [21], [22] . 49
E.2 Calculation. 50
Annex F (informative) Protocol for spiking of solid matrices . 51
Bibliography . 52

European foreword
This document (CEN/TS 16800:2020) has been prepared by Technical Committee CEN/TC 444
“Environmental characterization of solid matrices”, the secretariat of which is held by NEN.
Attention is drawn to the possibility that some of the elements of this document may be the subject of
patent rights. CEN shall not be held responsible for identifying any or all such patent rights.
This document supersedes CEN/TS 16800:2015.
The main changes compared to the previous edition are listed below:
— the scope has been extended from water only to water and environmental solid matrices, thus the
document has been modified accordingly;
— a protocol for spiking of solid matrices has been added in an informative annex.
According to the CEN/CENELEC Internal Regulations, the national standards organisations of the
following countries are bound to announce this Technical Specification: Austria, Belgium, Bulgaria,
Croatia, Cyprus, Czech Republic, Denmark, Estonia, Finland, France, Germany, Greece, Hungary, Iceland,
Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland, Portugal, Republic of
North Macedonia, Romania, Serbia, Slovakia, Slovenia, Spain, Sweden, Switzerland, Turkey and the United
Kingdom.
Introduction
Environmental monitoring of chemical substances is increasingly carried out within a European
framework, and there is concern about the comparability of data at the European level. Methods used for
the monitoring of substances with recent interest have often not been properly validated either in-house
(i.e. within a single laboratory) or at the international level.
These issues may be addressed by adopting a harmonized approach towards method development and
validation. The main objective of this document is to provide a common European approach to the
validation of chemical methods for the respective monitoring of chemical substances in a broad range of
matrices. Although the development of this approach was triggered by the needs for monitoring of
emerging pollutants, it is of general nature and can be applied to the measurement of the concentration
of a wide range of substances in a variety of matrices.
This guidance considers the different requirements for the level of method maturity and validation at
different stages of the investigation or regulation of chemical substances.
This protocol will guide the user through the following steps:
— classification of existing methods with respect to their status towards validation, and the selection of
the appropriate validation approach;
— development of a method to extend its application; for example, if a method for determining a
required target compound in a selected matrix is available, but is not suitable for the same compound
in a different matrix of interest;
— the validation procedures to be undertaken to effectively demonstrate the validation status of a
selected method according to the approach adopted.
To agree on the use of one method, or several similar methods, in a trans-border or a multi-metrological
context, and allow comparison of the results reported by several data producers on the same location
(water quality measured on both bank of the same river, or soil composition measured on both sides of a
border, or continuity of quality assessment of waste after measurement provider, e.g.), the procedure of
establishing the LOQ of the measurement method must be clearly published.
The LOQ may result of:
— statistical evaluation of repeated measurements of a blank sample or a sample with a low
concentration of the compound of interest (LOQ);
— experimental verification with a spike matched matrix that the LOQ meets accuracy validation
criteria (LOQ-V).
Many (national and international) standards currently contain in their scope a statement like “this
method is applicable from a concentration level of xx µg/l or yy mg/kg dry matter”, without any
statement how this concentration level was established. When the limit of quantification is evaluated
(LOQ) or verified (LOQ-V) using the procedure of this guideline, there is a possibility that it does not meet
the lower limit of the claimed range.
Also, the LOQ and LOQ-V might be different depending on the analytical method. Therefore, if criteria are
set to the LOQ of a method, it is necessary to clarify if LOQ or LOQ-V is meant.
1 Scope
This document describes an approach for the validation of physico-chemical analytical methods for
environmental solid matrices and water.
The guidance in this document addresses the initial description of the method and two different
validation approaches, in increasing order of complexity. These are:
a) method development, if the method is developed by the laboratory, or conditions of adoption, if the
method is a standardized protocol adopted by the laboratory;
b) validation at the level of single laboratories (within-laboratory validation);
c) method validation at the level of several laboratories (between-laboratory or inter-laboratory
validation), with a focus on methods that are sufficiently mature and robust to be applied not only by
a few expert laboratories but by laboratories operating at the routine level.
The concept is strictly hierarchical, i.e. a method shall fulfil all criteria of within-laboratory validation
before it can enter the validation protocol of the between-laboratory.
This document is applicable to the validation of a broad range of quantitative physico-chemical test
methods for the analysis of water (including drinking water, surface water, groundwater, waste water,
marine water), and of solid environmental matrices, such as soil, sludge, liquid and solid waste, sediment
and biota. It is intended for standardized protocols adopted by a laboratory, and either for test methods
aiming at substances that have recently become of interest or for test methods applying recently
developed technologies.
The minimal requirements that are indispensable for the characterization of the fitness for the intended
purpose of an analytical method are: selectivity, precision, trueness, performances characteristics and
measurement uncertainty. The aim of validation is to prove that these requirements are met.
In this document after the definitions (Clause 3) and description of the principles (Clause 4) a toolbox is
given describing the relevant performance characteristics in the validation process.
Clause 7 and 8 focus on the within laboratory validation process (V1) and Clause 9 on the interlaboratory
validation process (V2). Clause 7 and 8 describe largely the same processes, but differ in approach for
establishing the LOQ.
Reporting of the results of the validation studies is addressed in Clause 10.
2 Normative references
The following documents are referred to in the text in such a way that some or all of their content
constitutes requirements of this document. For dated references, only the edition cited applies. For
undated references, the latest edition of the referenced document (including any amendments) applies.
ISO/IEC Guide 99:2007, International vocabulary of metrology — Basic and general concepts and
associated terms (VIM)
3 Terms and definitions
For the purposes of this document, the terms and definitions given in ISO/IEC Guide 99:2007 (VIM) and
the following apply.
3.1
accepted reference value
value that serves as an agreed-upon reference for comparison, and which is derived as:
a) a theoretical or established value, based on scientific principles;
b) an assigned or certified value, based on experimental work of some national or international
organization;
c) a consensus or certified value, based on collaborative experimental work under the auspices of a
scientific or engineering group;
d) when a), b) and c) are not available, the expectation of the (measurable) quantity, i.e. the mean of a
specified population of measurements
[SOURCE: ISO 3534-2:2006, definition 3.2.7]
3.2
accuracy
closeness of agreement between a test result and the accepted reference value
Note 1 to entry: The term accuracy, when applied to a set of test results, involves a combination of random
components (usually expressed by a precision measure) and a common systematic error or bias component (usually
expressed by a measure for trueness).
Note 2 to entry: The technical term “accuracy” should not be confused with the term ‘trueness’ (see definition of
“trueness”).
[SOURCE: ISO 3534-2:2006, definition 3.3.1]
3.3
analyte
substance to be analysed (chemical species or physical parameter)
Note 1 to entry: The quantity of an analyte is the measurand (3.15).
3.4
bias
difference between the expectation of a test result or measurement result and a true value
Note 1 to entry: Bias is the total systematic error as contrasted to random error. There may be one or more
systematic error components contributing to the bias. A larger systematic difference from the accepted reference
value is reflected by a larger bias value.
Note 2 to entry: The bias of a measuring instrument is normally estimated by averaging the error of indication
over an appropriate number of repeated measurements. The error of indication is the: “indication of a measuring
instrument minus a true value of the corresponding input quantity”.
Note 3 to entry: In practice, accepted reference value is substituted for the true value.
[SOURCE: ISO 3534-2:2006, definition 3.3.2]
3.5
blank
sample or test scheme without the analyte known to produce the measured signal
Note 1 to entry: Use of various types of blanks enable assessment of which proportion of the measured signal is
attributable to the measurand and which proportion to other causes. Various types of blank are available (see
definition of Reagent Blank and Sample Blank).
3.6
calibration
operation that, under specified conditions, in a first step, establishes a relation between the quantity
values with measurement uncertainties provided by measurement standards and corresponding
indications with associated measurement uncertainties and, in a second step, uses this information to
establish a relation for obtaining a measurement result from an indication
Note 1 to entry: A calibration may be expressed by a statement, calibration function, calibration diagram,
calibration curve, or calibration table. In some cases, it may consist of an additive or multiplicative correction of the
indication with associated measurement uncertainty.
Note 2 to entry: Calibration should not be confused with adjustment of a measuring system, often mistakenly
called “self-calibration”, nor with verification of calibration.
[SOURCE: ISO/IEC Guide 99:2007, definition 2.39]
3.7
certified reference material
CRM
reference material, accompanied by documentation issued by an authoritative body and providing one
or more specified property values with associated uncertainties and traceabilities, using valid procedures
[SOURCE: ISO/IEC Guide 99:2007, definition 5.14]
3.8
fitness for purpose
degree to which data produced by a measurement process enables a user to make technically and
administratively correct decisions for a stated purpose
3.9
intermediate precision
precision under intermediate precision conditions
[SOURCE: ISO 3534-2:2006, definition 3.3.15]
3.10
intermediate precision conditions
conditions where test results or measurement results are obtained with the same method, on identical
test/measurement items in the same test or measurement facility, under some different operating
conditions
Note 1 to entry: There are four elements to the operating condition: time, calibration, operator and equipment.
[SOURCE: ISO 3534-2:2006, definition 3.3.16 and ISO 11352:2012, definition 3.10]
3.11
limit of detection
LOD
measured quantity value, obtained by a given measurement procedure, for which the probability of
falsely claiming the absence of a component in a material is β, given a probability α of falsely claiming its
presence
Note 1 to entry: IUPAC recommends default values for α and β equal to 0,05.
Note 2 to entry: The abbreviation LOD is sometimes used.
Note 3 to entry: The term “sensitivity” is discouraged for ‘detection limit’.
Note 4 to entry: The LOD is the lowest concentration of measurand in a sample that can be detected, but not
necessarily quantitated under the stated conditions of the test.
[SOURCE: ISO/IEC Guide 99:2007, definition 4.18]
3.12
limit of quantification
LOQ
lowest concentration of a measurand that can be determined with acceptable precision under the stated
conditions of the test
Note 1 to entry: as such defined, LOQ is based on evaluation of precision. This does not encompass neither any
eventual bias, nor laboratory measurement uncertainty at LOQ level.
3.13
Verified LOQ
LOQ-V
lowest concentration of a measurand that can be determined with acceptable accuracy under the stated
conditions of the test
Note 1 entry: LOQ-V is based on the check of a defined level of accuracy of the method at LOQ-V level. Bias and
precision have been considered to verify LOQ-V
3.14
reporting limit
RL
specific concentration at or above the limit of quantification that is reported to the client with a certain
degree of confidence
Note 1 to entry: The reporting limit is often defined on a project-specific basis. If the reporting limit is set below
the limit of quantification by the client, method modification is required
[SOURCE: ISO/TS 13530:2009, 4.4.7]
3.15
linearity
ability of the method to obtain test results proportional to the concentration of measurand
Note 1 to entry: The linear range is by inference the range of measurand concentrations over which the method
gives test results proportional to the concentration of the measurand.
[SOURCE: EURACHEM Guide]
3.16
measurand
quantity intended to be measured
Note 1 to entry: The specification of a measurand requires knowledge of the kind of quantity, description of the
state of the phenomenon, body, or substance carrying the quantity, including any relevant component, and the
chemical entities involved.
Note 2 to entry: In chemistry, “analyte”, or the name of a substance or compound, are terms sometimes used for
“measurand”. This usage is erroneous because these terms do not refer to quantities.
[SOURCE: ISO/IEC Guide 99:2007, definition 2.3]
3.17
measurement
process of experimentally obtaining one or more quantity values that can reasonably be attributed to a
quantity
[SOURCE: ISO/IEC Guide 99:2007, definition 2.1]
3.18
measurement uncertainty
non-negative parameter characterizing the dispersion of the quantity values being attributed to a
measurand, based on the information used
[SOURCE: ISO/IEC Guide 99:2007, definition 2.26]
3.19
outlier
member of a set of values which is inconsistent with the other members of that set
Note 1 to entry: ISO 5725-2 specifies the statistical tests and the significance level to be used to identify outliers
in trueness and precision experiments.
[SOURCE: ISO 5725-1:1994, definition 3.21]
3.20
precision
closeness of agreement between independent test results obtained under stipulated conditions
Note 1 to entry: Precision depends only on the distribution of random errors and does not relate to the true value
or the specified value.
Note 2 to entry: The measure of precision is usually expressed in terms of imprecision and computed as a
standard deviation of the test results. Less precision is reflected by a larger standard deviation.
Note 3 to entry: “Independent test results” means results obtained in a manner not influenced by any previous
result on the same or similar test object. Quantitative measures of precision depend critically on the stipulated
conditions. Repeatability and reproducibility conditions are particular sets of extreme conditions.
[SOURCE: ISO 3534-2:2006, definition 3.3.4]
3.21
proficiency testing
evaluation of participant performance against pre-established criteria by means of interlaboratory
comparisons
[SOURCE: EN ISO/IEC 17043:2010, definition 3.7]
3.22
quality assurance
part of quality management focused on providing confidence that quality requirements will be fulfilled
Note 1 to entry: A major part of quality assurance is quality control.
[SOURCE: EN ISO 9000:2015, definition 3.3.6]
3.23
quality control
part of quality management focused on fulfilling quality requirements
[SOURCE: EN ISO 9000:2015, definition 3.3.7]
3.24
quantity
property of a phenomenon, body, or substance, where the property has a magnitude that can be
expressed as a number and a reference
[SOURCE: ISO/IEC Guide 99:2007, definition 1.1]
3.25
working range
interval, being experimentally established and statistically proved by the calibration of the method,
between the lowest and highest quantity possibly measured by the method
Note 1 to entry: The lowest possible limit of a working range is the limit of quantification of an analytical method.
3.26
reagent blank
all reagents used during the analytical process (including solvents used for extraction or dissolution) are
analysed in isolation in order to check whether they contribute to the measurement signal
Note 1 to entry: The measurement signal arising from the measurand can then be corrected accordingly.
3.27
Analytical recovery
extent to which a known, added quantity of analyte (3.3) in a sample can be measured by an analytical
system
Note 1 to entry: Recovery is calculated from the difference between results obtained from a spiked and an
unspiked aliquot of sample and is usually expressed as a percentage.
[SOURCE: Adapted from ISO 5667-14:2014, definition 3.8]
3.28
reference material
RM
material, sufficiently homogeneous and stable with respect to one or more specified properties, which
has been established to be fit for its intended use in a measurement process
Note 1 to entry: RM is a generic term.
Note 2 to entry: Properties can be quantitative or qualitative, e.g. identity of substances or species.
Note 3 to entry: Uses may include the calibration of a measurement system, assessment of a measurement
procedure, assigning values to other materials, and quality control.
Note 4 to entry: ISO/IEC Guide 99:2007[1] has an analogous definition (5.13), but restricts the term
“measurement” to apply to quantitative values. However, Note 3 of ISO/IEC Guide 99:2007, 5.13 (VIM), specifically
includes qualitative properties, called “nominal properties”.
[SOURCE: ISO Guide 30:2015, definition 2.1]
3.29
repeatability
precision under repeatability conditions, i.e. conditions where independent test results are obtained with
the same method on identical test items in the same laboratory by the same operator using the same
equipment within short intervals of time
[SOURCE: ISO 3534-2:2006, combination of definition 3.3.5 and definition 3.3.6]
3.30
reproducibility
precision under reproducibility conditions, i.e. conditions where test results are obtained with the same
method on identical test items in different laboratories with different operators using different
equipment
[SOURCE: ISO 3534-2:2006, combination of definition 3.3.10 and definition 3.3.11]
3.31
residual
difference between the observed response and that predicted by a calibration function
3.32
robustness
measure of capacity of a procedure to remain unaffected by small, but deliberate variations in method
parameters and provides an indication of its reliability during normal usage
3.33
sample
totality of a homogeneous analysis material with an identical composition or quality (similar to term
batch)
[SOURCE: ISO/TS 20612:2007, definition 3.3]
3.34
blank sample
matrix with no measurand
Note 1 to entry: They are difficult to obtain but such materials are necessary to give a realistic estimate of
interferences that would be encountered in the analysis of test samples.
3.35
selectivity
ability of a method to determine accurately and specifically the measurand of interest in the presence of
other components in a sample matrix under the stated conditions of the test
3.36
sensitivity
change in the response of a measurand divided by the corresponding change in the stimulus
3.37
traceability
property of a measurement result whereby the result can be related to a reference through a documented
unbroken chain of calibrations, each contributing to the measurement uncertainty
[SOURCE: ISO/IEC Guide 99:2007, definition 2.41]
3.38
trueness
closeness of agreement between the average value obtained from a large series of test results and an
accepted reference value
Note 1 to entry: In the present document, trueness will be expressed in terms of bias.
Note 2 to entry: Trueness should not be confused with the term ‘accuracy’ (see definition of “accuracy”).
3.39
validation
verification, where the specified requirements are adequate for an intended use
Note 1 to entry: This process is used to asses that a method is fit for its intended purpose. It includes:
— establishing the performance characteristics, advantages and limitations of a method and the
identification of the influences which may change these characteristics, and the extent of such changes;
— a comprehensive evaluation of the outcome of this process with respect to the fitness for purpose of the
method.
[SOURCE: ISO/IEC Guide 99:2007, definition 2.45]
3.40
verification
provision of objective evidence that a given item fulfils specified requirements
EXAMPLE 1 Confirmation that a given reference material as claimed is homogeneous for the quantity value and
measurement procedure concerned, down to a measurement portion having a mass of 10 mg.
EXAMPLE 2 Confirmation that performance properties or legal requirements of a measuring system are
achieved.
EXAMPLE 3 Confirmation that a target measurement uncertainty can be met.
Note 1 to entry: When applicable, measurement uncertainty should be taken into consideration.
Note 2 to entry: The item may be, e.g. a process, measurement procedure, material, compound, or measuring
system.
Note 3 to entry: The specified requirements may be, e.g. that a manufacturer's specifications are met.”
[SOURCE: ISO/IEC Guide 99:2007, definition 2.44]
3.41
application range
range of concentrations routinely measured by a method
[SOURCE: ISO 6107-2:2006, 8]
4 Principle
The goal of any validation activity is to demonstrate the applicability of the method to the intended
purpose for which it is to be used. Any laboratory performing analytical measurement shall provide
reliable and comparable data when carried out on similar samples, whatever the modifications of the
operating conditions, e.g. operators. This fitness-for-purpose shall be demonstrated through
experimental, well documented evidences. This comprises a set of general principles and criteria that are
applicable to physico-chemical test methods.
The requirements for methods used for determination of chemical measurands depend on:
— the extent of the intended or requested measurement activity; and
— the potential of the available methods that may be used for monitoring a specific substance (group).
In some cases, fully developed methods usable by routine laboratories may already exist. In the case of
new substances or newly developed technics there will be a lack of information on the extent to which
the methods have been fully developed and validated, which is concretely a default of publicity of the
performance characteristics of the method, and/or the way used to establish these performances.
To cover most eventualities, two distinct (and hierarchical) levels of method validation, V1 (7, 8), resp.
V2 (9), are described in this document. The validation levels are organized as shown in Figure 1.
Performance characteristics relevant for the demonstration of the fitness-for-purpose of a method are,
among others, selectivity, sensitivity, robustness, application range, measurement uncertainty. They
constitute the toolbox of the validation process, where method characteristics and their quantification
protocols are selected for the demonstration.
Figure 1 — Flow chart of a method validation
5 Characterization toolbox: performance characteristics
5.1 Introduction
An analytical method is characterized by performance criteria, limitations where existing, and influences
that may change them. The usual performance characteristics are:
— Selectivity (3.35);
— Sensitivity (3.36);
— Robustness (3.32);
— Trueness (3.37), bias (3.4) and method recovery (3.27), if applicable;
— Precision (3.20);
— Limit values: detection limit (3.11), quantification limit (3.12), verified quantification limit (3.13) and
if necessary, reporting limit (3.14);
— Calibration (3.6);
— Application range (3.40);
— Measurement uncertainty (3.18).
These characteristics should be determined for all validation levels. The experimental design may vary
depending on the level. The suitability of the method documented according to Annex A is investigated
on the whole scope. When an extension of the scope of a validated test method is to be investigated,
validation shall entirely be carried out on the additional parts of the new scope.
The design of experiment at validation level V1 and at level V2 is the same. This design of experiment is
detailed in Table A.2.
5.2 Characteristics
5.2.1 Selectivity
Selectivity should be taken into consideration, albeit more in qualitative terms of the attention that has
been paid to possible interferences than in terms of quantification.
Selectivity is a performance characteristic that is difficult to quantify. It is necessary to establish that the
signal produced by the measurement system, or other measured property, is unambiguously attributed
to the measurand, and not produced by accident or coincidence or due to the presence of chemically or
physically similar compounds. The selectivity of a method can usually be investigated by studying its
ability to measure the substance of interest compared to selected interferences which have been
introduced in the sample (i.e. those interferences thought likely to be present in samples or which from
expert knowledge are known to be likely interferences for the method). Another indirect way is checking
the bias of a method (e.g. by analysing a CRM). However, often a CRM is not available for chemicals of
recent interest, and a possible interference should also be present in the CRM. If a bias consistent with
the fitness for purpose of the method cannot be demonstrated by analysis of CRMs that also contain
representative amounts of interfering compounds, then the method is considered as selective.
Analyses of spiked and non-spiked samples with and without interfering substances allow to distinguish
between matrix effects (change of sensitivity) and background interferences.
5.2.2 Sensitivity
Sensitivity (3.35) is related to the response of a measurand along with the change of its concentration in
the sample. Sensitivity should be established via the calibration function of the method.
5.2.3 Robustness
Robustness for routine use can be regarded as multiple:
a) robustness toward sample variation in one laboratory (matrix);
b) robustness towards small variations in environmental and/or operational conditions (3.32);
c) robustness toward variability in implementation by several different laboratories.
Robustness of the method toward sample variation shall be investigated during method development, in
any case before any collaborative trials. This approach helps also in saving costs by avoiding running
costly trials on insufficiently investigated methods.
For intra-laboratory validation, only a) and b) shall be considered.
The capacity of an analytical method to remain unaffected by small variations in environmental and/or
operational conditions provides an indication of its reliability during normal usage. This can be tested
using a systematic set of experiments that introduce small but deliberate changes to the experimental
conditions of the method, and by observing (either in a qualitative or quantitative way) how these
changes affect the final result by determining the relative standard deviations of e.g. the spike sample.
About the deliberate changes that are introduced, these can be different instrumental settings, reagents,
materials, amounts of sample material, exposure times, etc. Eventually, this approach should provide
information about the most critical conditions that affect the performance and reliability of the method.
To examine the effect of the variation in the environmental conditions on the results, a “factorial design”
approach could be applied as described in Annex F. The advantage of this approach is that information
can be provided on which environmental/operational conditions significantly affect the results.
For interlaboratory validation, robustness as in c) should be considered. Organization of such trials is
described in Annex C.
5.2.4 Trueness
5.2.4.1 Bias
The trueness of the method shall be evaluated to investigate the potential for bias in the method.
x
The method bias b can be estimated by comparing the mean of analytical results (with uncertainty
u x u
) with the assigned value (uncertainty )
x av av
b xx−
(1)
av
The standard uncertainty of the bias results from the combination of the above-mentioned uncertainties:
2 2
u uu+ (2)
b x av
With the coverage factor k = 2 (for about 95 % confidence) the expanded uncertainty is
Uu2×
(3)
bb
=
=
=
As long as the bias b is lower than its expanded uncertainty U the bias may be regarded as not significant.
b
5.2.4.2 Method recovery
To evaluate a method recovery, an accepted reference value (often referred to as “conventional true
value”) is essential. Ideally, the accepted reference value is established for a so-called certified reference
material (CRM). For pollutants with recent interest that are to be measured using methods validated at
the V1 level, the availability of CRMs is unlikely. In the absence of a suitable CRM, consensus mean, or
median values of ring test samples are often used as an estimate of accepted reference values. At the V1-
level, neither CRMs nor ring test results may be available.
As an alternative, spiking a sample with a known amount of measurand and analysing the sample before
and after spiking offers a means of determining recovery. The recovery shall be calculated on at least 2
levels, as described in e.g. ISO 11352:2012, 8.3.2 or 8.3.4.
NOTE 1 The spiking level can influence the bias of the method when using this approach. Lower spike
concentrations might give a larger relative bias.
A homogeneous bat
...