Method Development and GLP Bioanalytical Method Validation

Chemical and biological drug concentration measurement and their associated metabolites are key to pivotal nonclinical toxicokinetic (TK) and pharmacokinetic (PK) studies and Phase I-III clinical trials. The results from well-characterized, appropriately validated, and documented methods ensure reliable data to support regulatory decisions for your program.

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Bioanalytical Method Development

Bioanalytical method development for both small and large molecules must carefully define the design, operating conditions, limitations, and suitability of a method that is ultimately optimized for method validation. Method development involves finding the best procedures and conditions for extracting and detecting the analyte and/or its metabolites, understanding that each bioanalytical technique has its own characteristics and will vary from analyte to analyte. Our scientists are constantly developing and validating methods for both small molecules and biologics across many technology platforms including LC/MS/MS, LC/HRMS, ligand binding assays, automated ELISA systems, Gyrolab, RIA, SPA, MSD, Flow Cytometry Assays, qPCR and ddPCR. The network of bioanalytical chemists within Charles River provides an enormous knowledge bank to pull from for the most difficult of assays. We are also the world’s leading CRO in microsampling.

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What Our Client Says About Us?

"Working with the analytical chemistry group, and specifically Maryann Nolan, Research Scientist, Mattawan, MI, has gone smoothly for over three years now. Maryann is extremely knowledgeable, organized, and always delivers the data on time. She is an excellent communicator and keeps us informed if there are delays or issues with the data and she has helped with investigations when we have had unexpected findings or results to resolve quickly. She delivers reports on time, and they are always of high quality. I cannot say enough great things about how wonderful it is to work with Maryann, particularly on more challenging moieties like oligonucleotides."

- Dinah Misner, PhD, DABT, DSP
Senior Director | Aligos Therapeutics Inc.

Bioanalytical Method Validation

It is essential to employ well-characterized fully validated bioanalytical methods to ensure reliable results to make those critical go/no-go decisions along the drug development journey. Each bioanalytical method is validated according to applicable health agency guidance documents by our scientists so that it is appropriate for the intended meaning of a study and yields robust data for specificity, precision, and accuracy. From preclinical studies to the late-phase clinical studies, a bioanalytical method may undergo modifications including the lower limit of quantification, additional metabolites, adjustments to minimize interference (particularly in a disease-state matrix), or a new biological matrix. Experience is key to minimizing delays. Whether your interest is in small or large molecules (e.g., antibody-drug conjugates, biosimilars, oligonucleotides, proteins, vaccines, gene therapy) our dedicated global team of some of the best bioanalytical chemists will ensure that your method validation and subsequent drug quantitation studies meet your timelines and regulatory requirements.

Bioanalytical Method Validation Guidelines

Different types or levels of validation may be employed including full method validation, partial validation, or cross-validation. All three method validation types are well-defined in the draft ICH Bioanalytical Method Validation Guidelines.

  • Full Bioanalytical Method Validation Elements

    Chromatographic Methods

    • Selectivity
    • Specificity
    • Matrix effect
    • Calibration curve
    • Range - LLOQ to ULOQ
    • Accuracy
    • Precision
    • Carry-over
    • Dilution integrity
    • Stability
    • Reinjection reproducibility

    Ligand Binding Assays (LBAs)

    • Selectivity
    • Specificity
    • Calibration curve
    • Range - LLOQ to ULOQ
    • Accuracy
    • Precision
    • Carry-over
    • Dilution linearity
    • Parallelism
    • Stability
  • Partial Bioanalytical Method Validation

    Modifications to a fully validated method may be evaluated by the conduct of partial validation, ranging from as little as one within-run accuracy and precision determination to a nearly full validation.

  • Cross-Bioanalytical Method Validation

    Cross-validation should be applied when data are obtained from different methods within or across studies, or when data are obtained within a study from different laboratories applying the same method, where a comparison of those data is needed.

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Frequently Asked Questions (FAQs): Bioanalytical Method Validation

  • Where can I find guidelines for bioanalytical method validation?

    International guidelines issued by the FDA and EMA provide recommendations for bioanalytical method validations for chemical and biological drug quantification and the application of those methods in the analysis of study samples. It is critical that the bioanalytical methods are validated in accordance with the appropriate guidelines, based on where the drug application will be submitted for approval. 

  • What parameters should be optimized for method development?

    To ensure a bioanalytical method is suitable for validation, method development should include the optimization of critical reagents, calibration curve, quality control samples, selectivity, specificity, sensitivity, accuracy, precision, recovery, stability of analyte in matrix, and, for ligand binding assays, minimum required dilution (MRD).

  • What is matrix effect and how do I evaluate it during method validation?

    A matrix effect is the alteration of the analyte response due to interfering components in a sample matrix. Different independent sources or lots of matrix should be evaluated during the method validation phase by analyzing replicates of low and high QCs each prepared from at least six different sources or lots.

  • What is bioanalytical dilution integrity and dilution linearity?

    If a sample dilution procedure is required, it must be confirmed to not impact measuring the accuracy and precision of the analyte of interest; so, dilution QCs are prepared with analyte concentrations that are greater than the ULOQ and then diluted with blank matrix. The dilution linearity experiment will demonstrate the high concentrations of the analyte of interest can be accurately measured by diluting into the assay’s quantitative range and multiplying the measured concentration by the dilution factor.