Host Cell DNA Assay
Very strict quality and product-safety regulations are entailed on products isolated from biological sources. These products are potentially contaminated by nucleic acids derived from the host cell system or other components used during cultivation and purification. Since most of the biological pharmaceuticals are delivered directly into the blood stream of the patients, there is a considerable risk due to oncogenic or other pathogenic effects of the nucleic acid, especially the more stable DNA. Therefore, the quantification of residual host cell or process related DNA in purified product is a pre-requisite for product release of biopharmaceuticals. |
 Threshold system: Total DNA Assay |
Regulatory authorities in Europe (EMEA), USA (FDA) and Japan (MHW) in the recent years have recognised that the risk of DNA from continuous mammalian cell lines (CCLs) was partially overestimated. Nevertheless, a limit for DNA impurities of 100 pg/dose is often discussed. In this context Glycotope Biotechnology offers a GCLP (Good Control Laboratory Practice) conform quantification of residual DNA in pharmaceutical proteins with high sensitivity (0.01 ppm).
Using the Threshold Total DNA Assay™ (Molecular Devices) it is possible to perform the quantitation of residual DNA with high precision and sensitivity (up to 4 pg/ml). In the Threshold Total DNA Assay the detection of DNA is based on the formation of an complex of single-stranded DNA, the single-stranded DNA binding protein (SSB) from E. coli, a monoclonal antibody, and streptavidin.
For establishment of routine testing an evaluation of the test article has to be done. Some low molecular weight components may inhibit binding and subsequent enzymatic reagents (list available on request). Furthermore, it was shown that some proteins inhibit the DNA assay. The matrix effects like inhibiting low molecular weight components or proteins have to be checked for the sample. Via dilution, concentration and spike recovery experiments it is checked, what kind of pre-treatment procedure is required (feasibility study).
Once the pre-treatment procedure is established, a validation study has to be performed to show accuracy, precision, limit of detection, and limit of quantification within the sample matrix according to the ICH guideline Q3B. The robustness of the assay can be tested by investigation of different batches of the sample.
Results of routine analysis are provided within 10 working days.
Glycotope Biotechnology is also prepared to offer DNA quantitation specific for host cell DNA on basis of Slot-Blot/Dot-Blot method. For special purposes it is possible to develop suitable primers and standards for quantitation of highly specific targets.
For further information and inquiries, please contact us (Dr. Ansgar Hofmann, phone +49 6221 9105-51, e-mail: mob@glycotope-bt.com)
Supplementary Information
PDF documents (need Acrobat Reader):
Total DNA assay summary
List of inhibitory substances
Test article supplementary information form (can be filled with Acrobat Reader)