Assays validation protocol

The accuracy of the method was evaluated by assaying six independently prepared solutions of IB-367 against two standard solutions of the same lot as external standards. The mean of 102.3% met the criteria set in validation protocol (97 to 103%). 

Execution of the method validation protocol should be carefully planned to optimize the resources and time required to complete the full validation study. For example, in the validation of an assay method, linearity and accuracy may be validated at the same time as both experiments can use the same standard solutions. A normal validation protocol should contain the following contents at a minimum ... 

High-performance liquid chromatographic methods are the most common form of analytical technique used to support drug product registration. An example of a validation protocol for an HPLC assay and related compounds method is provided in Example 2. 

TABLE 9-5. Range for Recovery Experiment for CU and Assay Method to Be Defined in Method Validation Protocol... 

Additionaf reading on the subject of assay validation and specific protocols can be found online at the NCGC web site Assay Guidance Manual (http //www.ncgc.nih.gov/guidance/index.html). 

Table 2 contains an example of validation target acceptance criteria for a stability-indicating HPLC method for the assay of an active drug substance and its related compounds. These criteria or others cited in the literature 22>23 can be used as a general guideline when considering acceptance criteria for a validation protocol. 

A validation protocol states the purpose of the validation method, the intended substance being tested, the definition of the reportable value, the validation approach, the specific directions for conducting the validation assays, the statistical approach for analyzing the resulting data, and the nonambig-uous acceptance criteria. The protocol should allow no room for ambiguity in the execution of the protocol or in the acceptance criteria. The content of a protocol is described next. There are many ways to format a protocol herein are suggestions for the content of a protocol. 

Once a new analytical method has been developed, it must be optimized and standardized to meet the purposes for which it was developed. Validation studies confirm that a new assay has met its required performance specifications, and must be done before the analytical procedure can become a routine laboratory protocol. There are multiple approaches to validating analytical methods. The most common is to compare results from the new method with results obtained using an established, or reference, assay. Alternatively, standard reference materials (e.g. standards certified by NIST) can be analyzed by the new method to demonstrate its accuracy. When available, authentic specimens should be tested and compared against a previously validated protocol. 

The process validation protocol is a preapproved written plan stating how the validation will be conducted and identifying acceptance criteria as well as sampling and assay requirements [33] and other testing details [3]. It is a prospective experimental plan that when executed produces documented evidence that the system has been validated [25]. The protocol defines the system to be validated, identifies operating variables and probable control parameters, and indicates the number of... 

An important aspect is that of data and model stability. The raw data, for example, assay data, need to be updated regularly as new results become available. This process is tedious and time consuming and once a valid protocol for data acquisition and validation has been established, it is suitable for automation. The automatic procedure requires extensive data integrity checks (e.g., do all data points have valid structures and experimental results, how to handle conflicting results from different experiments) and a formalized automated normahzation process. This involves answering questions such as how to handle isomers, do experimental in vivo results have precedence over in vitro results or should all results be shown, should more confidence be put in results from Good Laboratory Practice (GLP)-studies, and so on. The models are then automatically rebuilt using the updated data and auto-updated as was mentioned in the QSAR section. A model may be promoted to use in the production system if it passes some defined validation tests, specific for each model, which was also mentioned earlier. 

Two landmark articles published by H. Mark et al. and G. E. Ritchie et al. [30, 31] demonstrate how ICH Q2 principles can be applied when developing a near-infrared method. The first part of the series describes the method validation concepts as described in this section and proposes additional validation protocols. The second part focuses on the implementation of assay and content uniformity methods for solid dosage forms. Authors present at length how each criterion was validated and how they meet ICH and FDA requirements. For instance, repeatability was tested with the predictions of 13 repeats of tablets at three different concentration levels (80,100, and 120% w/w). In addition to the stated guidelines, authors used ASTM E1655-97 [12] to provide statistics analysis that is not directly provided in ICH Q2(R1). 

H. Mark, G. E. Ritchie, R. W. Roller, E. W. Ciurczak, C. Tso, and S. A. MacDonald, Validation of a Near-Infrared Transmission Spectroscopic Procedure. Part A. Validation Protocols, /. Pharm. Biomed. Anal, 28,251 (2002). G. E. Ritchie, R. W. Roller, E. W. Ciurczak, H. Mark, C. Tso, and S. A. MacDonald, Validation of a Near-Infrared Transmission Spectroscopic Procedure. Part B. Application to Alternate Content Uniformity and Release Assay Methods for Pharmaceutical Solid Dosage Forms, /. Pharm. Biomed. Anal, 29,159 (2002). M. Blanco, M. Bautista, and M. Alcala, API Determination by NIR Spectroscopy across Pharmaceutical Production Process, AAPS PharmSciTech, 9,1130 (2008). A. Peinado, J. Hammond, and A. Scott, Development, Validation and Transfer of a Near Infrared Method to Determine In-Line the End Point of a Fluidised Drying Process for Commercial Production Batches of an Approved Oral Solid Dose Pharmaceutical Product, /. Pharm. Biomed. Anal, 54,13 (2011). 

Before the performance of method transfer activities involving protocols and acceptance criteria, it was customary for a receiving laboratory to repeat some or all of the validation experiments. This laboratory was thereby deemed to be qualified when the results were demonstrated to meet the acceptance criteria described in the validation protocol. The choice of validation parameter(s) depends highly on the type of method being transferred. For example, content uniformity assays to determine consistency of product potency depend heavily on the method and system precision. As a second example, a determination of trace impurities in an API could not be reproduced between two sites if their instruments did not yield similar limits of detection and/or quantitation. A detailed discussion on the rational choice of validation parameters that would need to be repeated by the receiving laboratory is beyond the scope of this chapter. The reader is referred to the method validation chapters by Wood (in-process methods) and Clarke and Bretnall (API and drug product methods) for additional information on this subject. 

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