Advancing PCR bioanalysis
Shedding of gene therapy products
At TATAA Biocenter, we collaborate closely with you to custom-design assays that accurately detect your shed products in excreta, secreta, or skin swabs. Our highly sensitive assays are validated to be in line with regulatory requirements.
Shedding guided by FDA and EMA
Preclinical and clinical shedding
Digital PCR (dPCR) outperforms quantitative PCR for shedding. We develop, optimize, and validate highly sensitive dPCR assays for shedding assessments. The assay sensitivity is critical when dealing with shed nucleic acids from replication-incompetent viral vectors, often present in very low copy numbers. The matrices in which these nucleic acids are found pose additional challenges, including DNA-degrading enzymes, PCR-inhibitory substances, and high levels of genetic background from endogenous flora and environmental microorganisms. We routinely overcome these challenges.
Recommendations and analysis of shedding studies by FDA and EMA
At TATAA Biocenter, we adhere to FDA and EMA guidelines, which mandate the study of preclinical and clinical shedding to mitigate the risk of transmission to untreated individuals. Our assays are designed to offer the sensitivity needed to detect shedding across various matrices, such as excreta, secreta, or the skin. The selection of these matrices is based on the nature of the product, its tropism, and the route of administration.
Immunological considerations
Pre-existing immunity, or immune priming during repeated administrations, can significantly affect drug efficacy, necessitating immunity testing to ensure reliable data. Using only microliters of plasma or blood samples, we use Olink Target 48 Cytokine panels to monitor the activation of the innate immune response through absolute quantification of key cytokines.
Shedding assay design
Extraction
The extraction method must be evaluated for each matrix and test item by spiking in targets to assess recovery after sampling, handling, storage, and extraction, given that the test item may exist in very low copy numbers.
Sample management, QC, and QA
Our regulated sample management processes include a Laboratory Information Management System (LIMS) for full traceability of samples and reagents, and a purpose-built laboratory to mitigate contamination risks and ensure the highest sensitivity possible. All data undergoes 100% QC, and QA according to regulatory requirements.
Assay development
We collaborate with you to design target-specific primers and probes and validate them for their specific context of use (COU), GLP, or GCLP. We conduct tests to ensure that the PCR’s sensitivity meets the stringent requirements for shedding assays and that the matrix does not interfere with the PCR.
Collaboration, reporting, and data transfer
Our principal investigators work closely with our sponsors, ensuring transparency in the process and project progression. Throughout the assay design, we keep sponsors informed of optimization steps, viewing ourselves as your extended workbench of experts. We customize the report to align with your downstream processes.
Viral shedding assays presents unique challenges. The analytes released are typically found in very low quantities, and the matrices involved in shedding assays often contain significant levels of PCR inhibitory agents and endogenous DNA. Our shedding services The FDA requires
Shedding assays require optimized extraction protocols tailored for each sample type and an accurate qPCR/dPCR assay with the necessary sensitivity and specificity. This ensures that shed products are detected and the risk of transmission to a third party is mitigated.
The Olink Target 48 panels provide both absolute and relative quantification. These panels include cytokines and other immune-related proteins, making them ideal for monitoring immune responses. They are perfect for assessing desired immune reactions in vaccines and CAR-T therapies, as
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