ACCELERATE PRODUCTION
Host cell DNA residual assays
We offer PCR-based host cell DNA residual assays with short lead times to accelerate process decisions.
Host cell impurities
DNA from the production host cell increases the risk of infection and oncogenicity. Therefore, safety testing for host cell DNA residuals is required for each batch.
Quality assessment of gene therapy products involves evaluating impurities not intended to be part of the final product. These impurities can be categorized as cellular impurities or process-related impurities, encompassing various elements such as reagents, media components, host cell components, residual DNA, viral vectors, vector-associated impurities, CRISPR-Cas9 ribonucleoproteins (RNPs), RNP-associated impurities, and other materials associated with viral production.
The risks associated with host cell DNA are potential infection and oncogenicity risks. The rep gene, utilized in vector production, possesses helicase and endonuclease activities that may increase mutation rates and promote vector integration in transduced cells.
Among DNA impurities, viral oncogenes represent a significant concern theoretically associated with an increased risk of oncogenesis. These genes are typically compact enough to fit within the viral capsid. Cell lines containing viral oncogenes are often used to produce AAV vectors. For instance, HEK 293 cells carry adenovirus E1, HEK 293T cells have adenovirus E1 and SV40 large T antigen, and HeLa cells harbor human papillomavirus E6 and E7. Plasmids containing the adenovirus E4 region, known for encoding proteins inhibiting the tumor suppressor p53 gene, are frequently used in triple transfection.
The risk to the human body increases as the length of residual host cell DNA fragments increases. Therefore, during the purification process, DNA is intentionally fragmented using nucleases. However, certain impurities may persist even after nuclease treatment, potentially because they are encapsulated within the capsid.
The primary goal of fragmentation is to disrupt the integrity of elements like oncogenes and eliminate infectious units such as integrated retroviruses and other functional sequences. It’s important to note that the AAV capsid shields the packaged nucleic acid from nuclease digestion, making removing or further reducing its size impossible.
Host cell DNA detection employs quantitative PCR methods like digital PCR. A representative segment, such as the 18S rRNA gene found in multiple copies within the HEK 293 cell genome, can serve as a target to identify host cell DNA. Ideally, we measure multiple amplicons of varying sizes to quantify a gene in more than one genome copy. Current guidelines recommend that residual cell-substrate DNA levels should not exceed ≤10 ng per dose, with a median DNA size of 200 bp or less.
Accelerate production with TATAA
We offer short lead times and fast analyses, allowing you to optimize the purification process, provide confidence in your products before GMP analysis, help with deviation control, and ensure trust in GMP results and safe product delivery.
Our well-equipped molecular analysis laboratory features automated platforms, a Laboratory Information Management System (LIMS), and high-throughput instruments for qPCR, dPCR, and Next-Generation Sequencing (NGS). Additionally, our facility includes a Biosafety Level 2 (BSL-2) laboratory that allows us to offer end-to-end solutions, from sample extraction to data transfer.
TATAA Biocenter can offer method development and validation tailored to your specific samples, from extraction to data transfer. We can assist you in measuring 18S rRNA, adenovirus E1, SV40 Large T antigen sequences, the E6/E7 genes, or any other genes you choose.
Stay in touch
Sign up for our newsletter
test
"*" indicates required fields
QUALITY ANALYSES
Accelerate with reliable results
With over 20 years of experience in nucleic acid analysis, we specialize in developing PCR-based assays that offer fast, accurate, and reliable quality testing for process optimization. The rapid results generated by PCR technology make it well-suited for assays on viable products, crucial for cell and gene therapies.
Vector copy number
Quantifying the transgene dosage, the presence of transgenes, the number of integrated vectors, or the number of non-integrated vector genomes in targeted cells provides essential information during production optimization.
Vector integrity
Ensuring consistent characterization of the packaged vector genome is crucial for production process optimizations and comparing results across production batches.
Pathogen detection
Various sequencing technologies provide a fast and reliable tool for evaluating sterility and detecting mycoplasma and endotoxins.
Integration site analysis
We evaluate the integration site specificity and transduction efficiency, assess safety, and prevent unintended effects on adjacent genes during integration.
Pharmacokinetics (PK) in gene therapies Pharmacokinetics for genes introduced by gene therapies is more challenging than those of finished small molecules. Our custom assays for biodistribution, pharmacokinetics (PK), and shedding. Ask a question Pharmacokinetics (PK) in gene therapies Pharmacokinetics for
About us
The power of our approach
Committed to quality, innovation, and fostering strong client relationships, we serve as your trusted partner to accelerate drug development. As a contract research organization, our mission is to push the boundaries of science and technology to generate accurate and reproducible data that shortens time-to-market.
Regulated laboratory environment
We are GLP accredited for qPCR, dPCR, and molecular biology, GCLP compliant, and ISO/IEC 17025 accredited. Our facility features a strict sample management process, a fully integrated LIMS system, backup for all vital systems, temperature and humidity control, and robust IT security.
Purpose-built PCR laboratory
Our purpose-built laboratory in Gothenburg, Sweden, is specifically designed for PCR. It has controlled air pressure, temporal separation, and biosafety cabinets to minimize contamination risk. This setup enables us to achieve the highest sensitivity and robustness required for validated assays. The lab is equipped with market-leading PCR and NGS instruments and advanced liquid handling systems.
Pioneers in assay development and validation
We are a team of 45 employees, with scientists at the forefront of assay development, optimization, and validation. Our team has co-authored key publications in the field, including Recommendations for Method Development and Validation of qPCR and dPCR Assays in Support of Cell and Gene Therapy Drug Development (AAPS J. 2024) and The MIQE Guidelines for qPCR and dPCR.
Flexible, client-centered solutions
We work closely with our clients to find tailored solutions, offering flexibility in sample types, test items, sample volume, and scalability. We prioritize transparency and proactive communication throughout each project, ensuring our clients are continuously updated on the progress.
contact us
Let's explore how we can support your drug development program
General Host cell DNA Page
"*" indicates required fields