Our experienced analytical development team works with customers through all stages of development, qualification and validation of the assays. If requested, we can transfer the final protocol to your labs (technology transfer) and organise comparability testing.
Our mission is to provide customers with high quality results, guarantee a quick turnaround time, address and solve any new challenges encountered and offer scientific support throughout the whole development process.
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ddPCR
We offer custom assay development, qualification and validation focused on analytical methods for quantification and detection of vector genomes and impurities (DNA, protein). We are qualified to analyse samples containing different virus vectors (e.g. AAVs, LVVs, AVs) which are three of the widely used vectors for delivering therapeutic genes to target cells and tissues.
ddPCR Sample Preparation
DNA / RNA extraction protocol development and qualification
Extraction of DNA and/or RNA can be a challenge, especially when various sample types and matrixes are used. Yeald, integrity and presence of inhibitory substances all effect quantification of the target DNA/RNA in the sample. The selection of most optimal extraction chemicals and sample preparation procedure, including nuclease treatment, thus need to be thoroughly planned and each step needs to be verified individually. We can develop custom DNA and RNA extraction protocols. Firstly, depending on the subsequent analytics (viral vector, hcDNA or pDNA quantitity), most appropriate protocols are selected and optimised. The extracted DNA and RNA is evaluated by different approaches offering a thorough overview of the DNA/RNA quality. Finally, the best performing protocol can be qualified and used in subsequent analyses.
Viral vector quantity
Viral vector copy number quantification by ddPCR
Viral vector copy number is determined by molecular methods, where digital PCR offers most accurate quantification. One of the crucial steps in addition to sample preparation, is suitable assay development and its qualification and/or validation. Depending on the target sequence and structure, more assays might be needed for reliable quantification. We can develop new custom assays and/or optimise existing assays (e.g. qPCR assays for use in ddPCR), evaluate their performance and qualify and/or validate best performing assays. Majority of our work is based on Bio-Rad’s ddPCR system, but we have access and experiences with other digital PCR platforms as well (Quantstudio 3D, Fluidigm, Qiacuity).
Residual DNA quantity
Host cell DNA and plasmid DNA quantification
Remainings of the cell line DNA and plasmid DNA in the final product is one of the most important safety concerns. In addition to different sample preparation procedures, also different approaches for hcDNA and pDNA quantification can be applied. We can develop new custom assays (e.g. for customer specific pDNA) or use existing protocols for quantification of hcDNA and pDNA with qPCR or ddPCR.
Host cell protein
Host cell protein (HCP) ELISA based quantification
In addition to DNA left overs from the upstream process, also proteins originating from cell line pose a big safety concern. Currently ELISA-based quantification is most widely spread and enables simple quantification of the HCP in different samples. We offer ELISA-based quantification of HCP and optimisation of the protocol.
Viral particle quantity and integrity
ELISA based quantification and electron microscopy
For wider insight into final sample quality, viral particles can be quantified and observed to determine presence of abnormalities. ELISA based quantification is a simple and fast system to quantify assembled viral particles, while negative staining electron microscopy offers unique opportunity to observe the particles themselves. We offer ELISA based viral particle quantification and, in collaboration with experts at NIB, TEM evaluation of full/empty ratio of particles, particle integrity, and possible impurities.
Applications
Our activities can support process development at different stages
- Sample analysis in support to pre-clinical and clinical studies in accordance with international testing guidelines and regulations,
- support in short-term and long-term stability studies,
- down-stream process optimization support,
- any other custom studies.
Infrastructure
We operate under Biosafety level 2 conditions and according to SIST EN ISO/IEC 17025:2017.
We are equipped with state-of-the-art equipment. In addition, our close connection with NIB and Life Science Park Mengeš allows us to use state-of-the-art institutional infrastructure. All devices and laboratory equipment are controlled and maintained in line with the requirements above.
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