What kind of testing should we be doing to evaluate our plasmid DNA quality and in what ways can quality affect vector manufacturing
This question is part of the following Ask The Expert session:
Plasmid DNA Production for Cell and Gene Therapy
Answered by:
Company: MilliporeSigma
Job Title: Associate Director, EMEA Segment Development for Cell and Gene Therapy
Answer
The quality and purity of plasmid DNA is of utmost importance for a successful transfection. Key watch outs are phenol, endotoxins and sodium chloride. Phenol and endotoxins can be harmful for the cells and salts can interfere with lipid complexing, which results in a decrease in transfection efficiency.
In order to determine the quality, one could assess (but not limited to):
- DNA concentration (OD 260 nm)
- DNA purity (OD 260/280 nm or UV-scan)
- Endotoxin (LAL test)
- Osmolality
- Residual genomic DNA (agarose gel electrophoreses or qPCR)
- Residual RNA (agarose gel electrophoresis)
- pH
- ccc monomer content (HPLC or via agarose gel electrophoresis)
With our BioReliance testing services, we offer a wide range of testing to helpyou assess the quality of your plasmid DNA that goes from Biosafety testing requirements (Sterility, Endotoxin, Host Cell Protein, Host Cell DNA, Sequencing, Plasmid conformation) to Product Characterization tests (Molecular Weight (ID) with Mass Spectrometry (MS), Purity – SEC/IEX/RP (depending on pDNA size), Concentration – A260/280)
This post is sponsored by MilliporeSigma.