BIOTECHNOLOGY
infectivity and immunogenicity. Guidance issued by the FDA and World Health Organization states that biotherapeutics developers “limit the amount of HCD for continuous non-tumorigenic cells to less than 10 ng/dose and the DNA size to below approximately 200 base pairs.” To comply with this guidance and
deliver a safe product with the appropriate therapeutic dose, manufacturers need to accurately quantify the amount of residual HCD. Here, qPCR, a common method for monitoring residual HCD, has drawbacks, including amplification bias, nonspecific signals, and lack of reproducibility. Furthermore, qPCR requires time- and labour-intensive sample extraction and calibration standards for quantification. Nanoplate digital PCR presents a
robust alternative with higher precision and sensitivity of detection at a lower template input range. Residual DNA (resDNA) quantification assays detect femtogram levels of HCD from CHO and
E.coli in a single reaction, even when PCR contaminants and other inhibitory reagents are present. Moreover, multicopy target assays ensure the results are not affected by the fragmentation of HCD. What’s particularly attractive is the digestion- and purification-free workflow, which reduces hands-on time.
MYCOPLASMA When producing adeno-associated virus (AAV)-mediated therapies, the use of live cell culture to grow AAVs opens up the possibility of potential mycoplasma contamination. Mycoplasma are difficult to detect, spread quickly, resistant to antibiotics and pathogenic to humans, rendering the entire production workflow vulnerable to failure. Until recently, qPCR was the “gold standard” for detecting process- or
Digital PCR based on microfluidic nanoplate technology takes quality control to the next level
product-related impurities, identifying mycoplasma in a day, but estimating its levels using a standard curve approach. Although this was considered an improvement over traditional culture-based tests, it is still not sensitive enough. Nanoplate dPCR offers a powerful
approach to rapidly identify contamination down to a single copy number and discriminate between live and dead microbes within complex gene therapy formulations. It supports advanced raw material and in-process testing, allowing more effective detection of microbial contamination and offering users 100% certainty about a product.
STERILITY TESTING Bioburden detection is critical for offsetting the risk of microbial contamination in the final product. However, delays in testing can cause
significant bottlenecks. Nanoplate dPCR speeds up bioburden detection and product release with higher throughput. Dedicated assays detect the total amount of viable microorganisms in or on a medical device, container, or component using multiplexed dPCR. Tis provides absolute quantification results and serves as an early warning system for possible production problems that could lead to inadequate sterilisation.
END-TO-END SOLUTIONS Te QIAcuity digital PCR system uses GMP-compliant software that meets the stringent requirements of U.S. FDA 21 CFR Part 11 regulations. Tis provides a secure environment for the maintenance, verification and tracking of all electronic records generated by the software during the biomanufacturing and QC process. Although today’s biomanufacturers are making great strides in discovering novel therapeutics, safety is still a concern and they remain vigilant in keeping their products free of contaminants and impurities. Technological advancements such as nanoplate digital PCR offer a big leap forward for safe and effective biomanufacturing.
The development of cell and gene therapies requires strict quality control
Dr Yvonne Collins is with Qiagen.
www.qiagen.com
www.scientistlive.com 35
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