Validation of immunoassays for bioanalysis: a pharmaceutical industry perspective

Area of Science:

• Bioanalytical Chemistry
• Pharmaceutical Analysis
• Immunology

Background:

• Immunoassays are vital bioanalytical methods for quantifying analytes via antigen-antibody reactions, widely used in pharmaceuticals for protein and biomarker analysis.
• Existing bioanalytical method validation guidelines offer limited specific recommendations for immunoassays.
• Immunoassays differ significantly from chromatographic assays in their underlying principles and validation requirements.

Purpose of the Study:

• To highlight key differences between immunoassays and chromatographic assays.
• To propose specific recommendations for the validation of immunoassay methods.
• To improve the quality and consistency of data generated by immunoassays.

Main Methods:

• Review of current bioanalytical method validation guidance.
• Comparative analysis of immunoassay and chromatographic assay characteristics.
• Development of recommendations for immunoassay-specific validation parameters.

Main Results:

• Immunoassay calibration curves are nonlinear, necessitating nonlinear curve fitting.
• Demonstrating immunoassay specificity is crucial due to the lack of prior analyte extraction.
• Recommended validation criteria for immunoassay accuracy and precision should be more lenient than for chromatographic assays.

Conclusions:

• The SFSTP confidence interval approach is recommended for evaluating total error in immunoassay validation.
• Implementing these recommendations can lead to more reliable and higher-quality immunoassay data.
• Standardized validation practices are essential for the consistent application of immunoassays in drug development and diagnostics.