10 Important Tips to Develop a Robust Test Method for GMP Testing of Biopharmaceuticals
Scientist
on
May 22, 2025
This blog post was written by Nuvalore, a service provider that offers innovative approaches, experienced teams and suitable analytical methods for all requirements.
With the increasing number of projects in the pipeline, many pharmaceutical companies are prioritizing the development of robust analytical platform test methods to enhance efficiency and accelerate outcomes. The establishment of these robust analytical methods is crucial, independent of test method scope. The methods can be developed for exploratory testing or as methods required for registration documents. Implementing platform methods minimizes the variety of mobile phases, columns, reagents and other components required. Additionally, the development of these methods facilitates smoother method transfers across affiliates, reduces investigation times following out-of-specification (OOS) or out-of-trend (OOT) results and offers regulatory flexibility. Following the tips below are not necessarily regulatory requirements but provide orientation enabling best laboratory practices, and basic knowledge in Quality by Design during method development is beneficial.
1. Evaluate existing projects and their existing methods
Compendial methods are excluded from the scope of this article due to their routine control. This article concentrates on non-compendial stability-indicating methods for antibody drug products. These methods include, but are not limited to:
- CE-SDS R/NR: Capillary Electrophoresis-Sodium Dodecyl Sulfate, Reduced/Non-Reduced
- iCiEF/cIEF: Imaged Capillary Isoelectric Focusing/Capillary Isoelectric Focusing
- SEC: Size-Exclusion Chromatography
- CEX: Cation-Exchange Chromatography
- HIC: Hydrophobic Interaction Chromatography
- HILIC: Hydrophilic Interaction Liquid Chromatography
The methods that should be used for optimization experiments are based on Analytical target profile (ATP). Once you have determined which methods are subject to optimization, make sure to list all existing projects and their corresponding methods within your company.
2. Factor collections
Quality by Design (QbD) and Design of Experiments (DoE) should be used to identify test method parameters that influence method performance. Factor collection should be carried out across different projects, making use of prior knowledge, product composition and molecule type. This phase of method development is crucial and should be carried out by a team experienced in drug development. Literature and published data should be reviewed in addition to any internal information. An Ishikawa diagram can be used during brainstorming sessions to illustrate the relationship between method parameters (factors) and the response, which represents method performance. It can also be used as initial risk assessment documentation.
3. Identify a reference standard that can be used across different projects
Determine the most suitable reference standard for evaluating the performance of the method across various projects. This involves selecting a reference standard that can be consistently applied to ensure reliable and comparable results of the developed platform method.
4. Identification of most important assay variables
The identification of critical assay parameters is an important step in the development of robust analytical methods. This process involves identifying and selecting parameters that significantly impact method performance. To select them in the bunch of factors (see tip 2), it may be beneficial to implement a scoring system that evaluates factors based on their impact on the method’s performance.
5. Screening for relevant factors using Design of Experiments approach
Design of Experiments is a systematic statistical methodology used to evaluate the influence of various factors on analytical method performance. The process begins with the screening of factors, which facilitates the application of either full factorial or fractional factorial designs. The fractional factorial approach allows for the efficient acquisition of information without the need to test all possible combinations. By employing screening DoE, researchers can determine the main effects of individual factors and identify interactions between them. The number of experiments required is contingent upon the complexity of the analytical method.
6. Test Method Optimization using DoE
When two or more factors are found to affect method performance, it is essential to refine the method through optimization. This involves, for instance, utilizing full factorial designs or response surface designs to systematically evaluate and enhance the test method. The refinement of test methods is carried out by adjusting the influential factors to achieve optimal performance. This process ensures that the method is robust and reliable. Robustness testing measures the insensitivity of the method to variations in parameters. It is the most effective way to assess the robustness studies during DoE experiments. Ideally, robustness and ruggedness should be tested before the project reaches the Stage 2 validation phase. This ensures that the method can consistently produce accurate and reliable results under varying conditions.
7. Verify the optimal assay conditions
To ensure the reliability of the established method, it is crucial to verify the optimal assay conditions. This involves repeating the optimal set of conditions to confirm consistency and accuracy. Verification can optionally be achieved through continuous monitoring of method performance. This ongoing tracking helps identify any deviations and ensures the method remains robust over time (See Tip 9).
8. Develop validation concept
When a new molecule is introduced, apply the newly developed platform method. If the platform method is unsuitable due to issues, proceed with method development to address these challenges.
To utilize the development data for future validation studies, finalize a comprehensive development report. This report should document all findings and optimizations, providing a solid foundation for subsequent validation phases.
Validation should be carried out under controlled conditions, adhering to guidelines for analysis, laboratory equipment, materials and other factors in line with ICH recommendations. Define acceptance criteria for accuracy, precision, linearity, specificity, detection and quantification limits, as well as the test method range. These criteria are crucial for ensuring the method’s robustness and reliability. The number of experiments and the effort required depend on the validation stage (Stage 1 or Stage 2). Each stage has specific requirements that must be met to ensure the method’s validity.
8. Track your platform method
Implement a trending tool to ensure that the method performance remains in a state of control throughout its entire lifecycle. This continuous monitoring is recommended to detect any deviations or trends that may indicate a loss of method robustness or reliability.
9. Document Thoroughly
Maintain detailed records of the method development process, including all findings and optimizations. This documentation supports regulatory submissions.
Please contact anamarija.curic@nuvalore.com from Nuvalore for more information about how you can improve your GMP test methods with nuvalore.