"The objective of validation of an analytical procedure is to demonstrate that it is suitable for its intended purpose" (International
Conference on Harmonisation Guideline Q2A).
"Methods validation is the process of demonstrating that analytical procedures are suitable for their intended use" (US
Food and Drug Administration Draft Guidance for Industry, 2000 ).
So, is your assay fit for the job ?
Validated analytical test methods are required by good manufacturing practice (GMP) regulations for products that have been
authorized for sale and almost certainly for late-stage trial clinical material.3 Also, some methods used during the pre-clinical
phase of drug development under good laboratory practice (GLP) regulations may also require validation.4
However, during the development of biopharmaceuticals, methods may be employed that may not need full validation — for example,
those used only for process validation or comparability studies. The various terms applied to the "not-quite-validation" of
such methods include "test characterization," "qualified method," and "validated for Phase I." Considerable confusion has
arisen over this topic, which has been the subject of several articles and numerous presentations at conferences. This article
also seeks to explain and clarify the situation.
In basic terms, a suitable method must be based on firm scientific principles; capable of providing the necessary sensitivity,
accuracy, precision, etc.; and capable of generating reliable results. During test development, we learn more about the ability
of a test to meet these requirements, and we decide whether the test is going to meet suitability standards. The key questions
to be answered are:
- What is the test objective?
- What performance characteristics must we look at?
- Is performance acceptable for the application?
- Can we have confidence in the results?
- How can we demonstrate assay validity?
The final "full" validation of a method, especially a biological assay, requires assembly of a significant amount of data
on the test's performance, so that the results produced can be subjected to statistical analysis and the variability of the
results documented. This can be an expensive, time-consuming procedure, and one would not want to go through the process until
it was absolutely necessary.
Regulatory Requirements and Guidelines
Current regulations with a bearing on this subject are listed in Table 1. The requirement to ensure adequate validation of
analytical methods originated in the US as a draft Subpart I of 21 CFR 211 in 1996. This stated "Methods Validation. The accuracy,
sensitivity, specificity and reproducibility of test methods used by a manufacturer shall be validated and documented. Such
validation and documentation shall be accomplished in accordance with Section 211.194(a)(2)."5 This latter section covers the laboratory records on test methods, both compendial and non-compendial. It specifically
requires that "The statement [of each method used in the testing of the sample] shall indicate the location of the data that
establish that the methods used in the testing of the sample meet proper standards of accuracy and reliability as applied
to the product tested. . . . The suitability of all testing methods used shall be verified under actual conditions of use."6
Table 1. Regulations and Guidances Which May Be Involved in Analytical Method Development and Validation