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Forensic Science Communications |
January 2005 – Volume
7 – Number 1 |
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Standards
and Guidelines |
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Validation of Analytical Methods
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Scientific Working Group for
the Analysis of Seized Drugs (SWGDRUG)
October 2003
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Introduction
| General
Validation Plan | Quality
Control | References
1. Introduction
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1.1.
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Validation is the confirmation by examination
and the provision of objective evidence that the particular
requirements for a specific intended use are fulfilled. There
are numerous documents that address the topic of validation,
but there are few validation protocols for methods specific
to seized drug analysis.
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1.2. |
An analytical scheme must be comprised of
validated methods that are appropriate for the analyte.
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1.2.1.
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The combinations of methods chosen for a particular
analytical scheme must identify the specific drug of interest,
preclude false positives, and minimize false negatives. |
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1.2.2.
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For quantification, the method should reliably
determine the amount of analyte present. |
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1.2.3.
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If validated methods are used from published
literature or another laboratory's protocols, then the
methods must be verified in each laboratory. |
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1.2.4.
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Verification should, at a minimum, demonstrate
that a representative set of reference materials has been
carried through the process and yielded the expected results.
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1.3.
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Each laboratory should determine whether its
current standard operating procedures have been validated,
verified, or require further validation and/or verification.
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1.4.
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All methods must be validated or verified
to demonstrate that they will perform in the normal operational
environment when used by people expected to use the methods
on casework.
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1.5.
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The entire validation and/or verification
process must be documented, and the documentation must be retained.
Documentation must include, but is not limited to, the following: |
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1.5.1.
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Personnel involved. |
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1.5.2.
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Dates. |
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1.5.3.
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Observations from the process. |
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1.5.4.
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Statement of conclusions and/or recommendations.
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1.5.5.
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Authorization-approval signature. |
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1.6.
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To meet these requirements, the Scientific
Working Group for the Analysis of Seized Drugs recommends that
laboratories follow the applicable provisions of the General
Validation Plan when validating seized drug analytical methods.
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2. General Validation Plan
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2.1.
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Purpose and/or Scope: This is an introductory
statement that will specify what is being tested, the purpose
of the testing, and the result(s) required for acceptance.
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2.1.1.
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Performance specification: A list
of specific objectives (e.g., trueness and precision) should
be determined prior to the validation process. |
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2.1.2.
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Process review: After completion
of the validation process, the objectives should be reviewed
to ensure that they have been satisfactorily met. |
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2.2.
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Analytical Method: State exactly the
method to be validated. It is essential that each step in
the method is demonstrated to perform satisfactorily. Steps
that constitute a method for the identification and/or quantification
of seized drugs may include the following:
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2.2.1.
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Visual characterization (e.g., macroscopic
examination). |
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2.2.2.
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Determination of quantity of sample, which may
include the following: |
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2.2.2.1.
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2.2.2.2.
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2.2.2.3.
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2.2.3.
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Sampling (e.g., representative or
random, dry, homogenized). |
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2.2.4.
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Sample preparation. |
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2.2.4.1.
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Extraction method. |
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2.2.4.2.
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2.2.4.3.
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2.2.4.4.
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2.2.4.5.
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Techniques for introducing sample
into instrumentation. |
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2.2.5.
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Instrumental parameters and specifications.
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2.2.5.1.
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List the instruments and equipment (e.g.,
balance and glassware) used.
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2.2.5.2. |
Instrument conditions. |
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2.2.6. |
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2.2.7.
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2.2.7.1.
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Equation(s) to be used. |
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2.2.7.2.
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Unit specification. |
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2.2.7.3.
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Number of measurements
required. |
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2.2.7.4. |
Reference values. |
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2.2.7.5. |
Significant figure
conventions. |
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2.2.7.6.
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Conditions for data
rejection. |
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2.2.7.7. |
Uncertainty determination.
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2.3.
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Validation Reference Materials: Appropriate
reference material(s) must be used for qualitative and quantitative
procedures. |
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2.4.
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Performance Characteristics |
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2.4.1.
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Selectivity: Assess the capability
of the method to identify/quantify the analyte(s) of interest,
whether pure or in a mixture. |
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2.4.2.
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Matrix effects: Assess the impact
of any interfering components and demonstrate that the method
works in the presence of substances that are commonly encountered
in seized drug samples (e.g., cutting agents, impurities, by-products,
precursors). |
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2.4.3.
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Recovery may be determined for quantitative
analysis. |
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2.4.4.
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Accuracy. |
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2.4.4.1.
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Precision (repeatability/reproducibility):
Determine the repeatability and reproducibility of all routine
methods. Conditions under which these determinations are made
must be specified. Reproducibility determination may be limited
to studies in the same laboratory. |
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2.1. |
2.4.4. |
2.4.4.1. |
2.4.4.1.1.
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Within the scope of the validation,
determine acceptable limits for repeatability and reproducibility. |
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2.4.4.1.2.
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For qualitative analysis, run the
qualitative method a minimum of ten times. |
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2.4.4.1.3.
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For quantitative analysis, run the
quantitative method a minimum of ten times. |
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2.4.4.1.4.
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Validation criteria for nonroutine
methods may differ from what is stated above. |
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2.4.4.2.
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Trueness must be determined for quantitative methods
to assess systematic error. Trueness can be assessed through
various methods such as the following: |
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2.4.4.2.1.
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Comparison of a method-generated
value for the reference material with its known value using
replicate measurements at different concentrations. |
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2.4.4.2.2.
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Performance of a standard-addition
method. |
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2.4.4.2.3.
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Comparison to proficiency test results. |
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2.4.4.2.4.
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Comparison with a different validated
analytical method. |
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2.4.5.
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Range: Determine the concentration or sample amount
limits for which the method is applicable. |
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2.4.5.1.
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Limit of detection must be determined for all
qualitative methods. |
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2.4.5.1.1.
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Determine the lowest amount of analyte
that will be detected and can be identified. (AOAC in Eurachem)
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2.4.5.1.2.
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The results obtained at the limit
of detection are not necessarily quantitatively accurate. |
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2.4.5.2.
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Limit of quantitation must be determined for all
quantitative methods. Determine the lowest concentration that
has an acceptable level of uncertainty. |
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2.4.5.3.
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Linearity must be determined for all quantitative
methods. |
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2.4.5.3.1.
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Determine the mathematical relationship
(calibration curve) that exists between concentration and response
over a selected range of concentrations. |
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2.4.5.3.2.
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The limit of quantitation effectively
forms the lower end of the working range. |
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2.4.5.3.3.
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Determine the level of acceptable
variation from the calibration curve at various concentrations.
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2.4.5.3.4.
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Determine the upper limits of the
working range. |
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2.4.6.
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Robustness must be determined for either qualitative
or quantitative methods. Alter the method parameters individually
and determine any changes to accuracy. |
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2.4.7.
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Ruggedness may be determined for either qualitative
or quantitative methods. Alter the analysts, instrumentation,
and environment to assess changes in accuracy. |
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2.5.
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Uncertainty: The contribution of random
and systematic errors to method result uncertainty must be assessed,
and the expanded uncertainty derived for quantitative methods. |
3. Quality Control
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Acceptance criteria for quality control parameters
should be adopted prior to implementing the method.
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4. References
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4.1.
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Enigma Analytical, Validating Analytical
Chemistry Methods: Enigma Analytical Training Course
(Version 2000-2001), Breckenridge, Colorado, 2000, pp 8-4,
8-5.
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4.2.
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Eurachem Working Group. Fitness for Purpose
of Analytical Methods: A Laboratory Guide to Method Validation
and Related Topics, Eurachem Guide, 1998. Available: www.eurachem.ul.pt/guides/valid.pdf.
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4.3.
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Department of Health and Human Services, Food
and Drug Administration. International Conference on Harmonisation:
Guideline on Validation of Analytical Procedures: Definitions
and Terminology. Federal Register, 60 FR 11260, Part
VIII (1995) pp. 11259-11262.
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